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HX00032735 


A  Text  Book 


OF 


Practical  Histology, 


WILLIAM    STIRLING. 

M.  D. 


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PRACTICAL     HISTOLOGY 


Frontispiece 


Mirvtem  BrOSf'litK 


;  I  THE  DETAILS  RLLED  IN . 


A    TEXT-BOOK 


OF 


PRACTICAL    HISTOLOGY 


WITH    OUTLINE    PLATES 


BY 


WILLIAM    STIRLING,    M.D.,   Sc.D.,   RR.S.E.  ' 

REGIUS    PROFESSOR    OF    THE     INSTITUTES     OF    MEDICINE     IN     THE     UNIVERSITY     OF    ABERDEEN 


SKilb  30  (!3«tlxitt  ;platcs,  1  Colourti)  |3lHtf,  ai>b  27  aJloob  6ngrafatngs 


PHILADELPHIA 
J.     B.     LIPPINCOTT     &     CO. 

LONDON  :    SMITH,    ELDER,    &   CO. 

i88i 

\All    rights     reservi:d\ 


<.. 


St  ^ 


TO 

CARL    LUDWIG 

PROFESSOR     OF     PHYSIOLOGY     IN     THE     UNIVERSITY     OF     I.EIPSIC 

AND 

L.    RANVIER 

PROFESSOR     OF     HISTOLOGY     IN     THE     COLLEGE     DE     FRANCE,     PARIS 

TO  WHOM  THE  AUTHOR  IS  INDEBTED  FOR  HIS  EARLIEST  TRAINING  IN  THE  TRUE 
METHOD  OF  MAKING  AND  RECORDING  ORIGINAL  OBSERVATIONS  IN  PHYSIOLOGY  AND 
HISTOLOGY,   HE,   WITH    HEARTFELT   PLEASURE,    DEDICATES   THIS   WORK. 

In    OFFERING    TO    THEM    THIS    TRIBUTE     OF    HIS    ADMIRATION     AND     REGARD,    THE 

author  desires  to  express  his  sense  of  the  inestimable  advantage  which 
they  have  conferred  on  physiology  and  histology. 

Their  labours,  characterised  as  they  have  been  by  enthusiasm  and  single- 
minded  DEVOTION  TO  THE  ADVANCEMENT  OF  SCIENCE,  HAVE  NOT  ONLY  ADDED 
GREATLY  TO  OUR  FUND  OF  KNOWLEDGE,  BUT  HAVE  ALSO  DEMONSTRATED  THE 
SUPERIORITY  OF  THAT  METHOD  OF  SCIENTIFIC  OBSERVATION  WHICH  CONSISTS  IN  A 
PATIENT   AND    INTELLIGENT   SCRUTINY   OF    NATURE. 

To  THIS  METHOD  THEY  HAVE  ATTRACTED  MANY  DISCIPLES  WHO  ACKNOWLEDGE 
THE  INCALCULABLE  VALUE  OF  THEIR  PERSONAL  INFLUENCE,  THE  CORDIALITY  WITH 
WHICH  THEY  HAVE  ENCOURAGED  AND  DIRECTED  THEM  IN  THEIR  PURSUIT  OF 
ORIGINAL  INVESTIGATIONS,  AND  THE  GENEROSITY  WITH  WHICH  THEY  HAVE  LAID 
OPEN  TO  THEM  THE  RICH  STORES  OF  KNOWLEDGE  ACCUMULATED  DURING  A  LIFE-LONG 
STUDY   OF   THE   PROBLEMS   OF   ANLMAL   STRUCTURE   AND    FUNCTION. 


PREFACE. 


The  ruKPOSE  of  this  work  is  twofold  :  first,  to  give  plain,  definite,  and  precise 
directions  for  the  preparation  and  examination  of  the  animal  tissues  ;  and,  secondly, 
to  ensure  that  the  student  executes  a  drawing  of  the  majority  of  the  microscopic 
specimens  which  he  makes  for  preservation.  For  this  purpose  a  series  of  Outline 
Plates  is  issued  with  the  text. 

The  author  has  endeavoured  to  give  a  faithful  account  of  the  methods  which  he 
has  found  to  be  most  useful  for  the  preparation  of  each  of  the  tissues  and  organs  of 
the  body  for  microscopic  purposes.  No  method  is  introduced  which  he  has  not 
found  from  repeated  trials  to  be  successful.  The  methods  described  are  those 
which,  after  nine  years'  experience  in  the  teaching  of  practical  histology,  he  has 
found  to  be  really  reliable. 

In  the  introductory  chapter  a  short  account  is  given  of  the  microscope  and  how 
to  use  it  ;  but  no  attempt  is  made  to  explain  the  optical  principles  on  which  it  is 
constructed,  as  the  author  believes  that  such  details  are  better  omitted  in  a  work 
like  the  present.  The  reader  is  therefore  referred  for  such  details  to  any  of  the 
numerous  works  upon  that  instrument.  The  student  is  advised  to  read  the  intro- 
ductory chapter  before  beginning  the  practical  portion  of  the  work. 

The  author,  from  long  experience  in  the  teaching  of  histology,  has  learnt  that 
the  reluctance  shown  by  students  in  making  drawings  of  their  microscopic  prepara- 
tions renders  it  advisable  to  facilitate  as  much  as  possible  this  method  of  exact 
representation.  This  he  does  by  giving  to  each  student  a  series  of  outline  plates, 
in  which  the  main  features  of  the  chief  microscopic  sections  are  indicated.  For  two 
sessions  he  has  done  this  with  his  classes  of  practical  histology  in  the  University 
of  Aberdeen,  and  he  finds  that  the  students  rapidly  acquire  the  art  of  fillin^T  in  the 
necessary  details.  At  the  end  of  the  course  of  histology,  therefore,  each  student  is 
in  possession  of  a  complete  set  of  microscopic  preparations,  which  illustrates  the 


viii  PREFACE. 

whole  range  of  normal  histology,  and  which  he  can  permanently  preserve  and  take 
away  with  him.  He  has  also  an  atlas  representing  the  details  of  the  more  impor- 
tant preparations ;  and,  lastly,  in  the  text-book  he  has  a  record  of  how  each 
specimen  was  prepared,  and  a  description  of  the  chief  points  to  be  observed  in 
each  microscopic  preparation.  It  is  to  be  remembered,  however,  that  no  mere 
instructions  or  directions  given  in  books  can  ever  supersede  that  form  of  informa- 
tion which  is  imparted  viva  voce  by  a  teacher,  and  which  is  elicited  by  the  difficulties 
that  meet  the  student  of  histology  at  the  beginning  of  his  course. 

He  is  indebted  to  Mr.  E.  A.  Schafer  for  the  use  of  several  of  the  woodcuts 
employed  in  the  introductory  chapter ;  and  for  fig.  3,  to  Messrs.  Baillicre,  Tindall, 
&  Cox.  The  outline  figures  were  drawn  by  the  author  from  the  microscopic 
specimens  with  which  each  student  is  supplied  or  makes  for  himself. 

He  has  also  to  express  his  sincere  thanks  to  Dr.  Robert  Lawson,  Deputy 
Commissioner  in  Lunacy,  and  to  Dr.  De  Burgh  Birch  for  revising  the  proof-sheets. 


WILLIAM    STIRLING. 


Physiological  Laboratory,  University  of  Aberdeen  : 
March,  1881. 


CONTENTS. 


INTRODUCTION. 


Histological  requisites 

The  use  of  the  microscope 

Artificial  illumination . 

Cleaning  of  the  objectives 

Method  of  focussing  an  object    . 

Rules  for  examining  an  object 

Preparation  of  a  simple  object     . 

Methods  of  applying  a  cover-glass  . 

Method  of  cleaning  a  cover-glass 

Measuring  cover-glasses 

Placing  a  section  on  a  slide 

Delineation  of  an  object         .         .         .     . 

Methods  of  studying  living  protoplasm 

Methods   of  applyitig  heat   to  miscroscopic 

objects  .      ■   .         .         .         .     . 

Methods  of  applying  vapours  to  objects 


PAGE 

xix 
xxi 
xxiii 
xxiii 
xxiii 
xxiv 
xxiv 
xxiv 

XXV 
XXV 

xxvi 
xxvi 
xxvii 

xxviii 
xxix 


Preliminary  preparation  of  the  tissues 
Methods  of  hardening  the  tissues 
Methods  of  softening  the  tissues 
Dissociation  of  the  tissues  . 
Methods  of  cutting  sections    . 
Microtomes        .... 
Methods  of  staining  the  tissues 
Aniline  dyes       .... 
Metallic  solutions  . 
Double  staining 
Treble-staining 
Staining  reagents 
Methods  of  mounting  sections 
Labelling  and  preserving  sections 
Methods  of  injecting  blood-vessels 
Methods  of  injecting  lymphatics 


PAGE 
XXX 

xxxi 

xxxiii 

xxxiv 

-xxxvi 

xxxvii 

xlii 

xliv 

xlv 

xlvi 

xlvii 

xlvii 

xlviii 

y.lix 

1 

Iv 


Blood  of  newt  or  fiog 

Human  blood 

Blood-crystals 

Epithelium 

Endothelium 

Cartilage 

Connective  tissue 

Bone 

Muscular  tissue    . 

Nerve-tissue    . 

Heart  and  blood-vessels 

Circulation  of  the  blood  . 

Larynx,  Trachea,  and  Lungs 

Lips  and  Tongue     . 

Tooth 

Salivary  glands 
Tonsils         .... 
Oesophagus  and  Stomach 
intestine      .... 
Liver       .... 
Pancreas      .... 
Lymphatic  system  . 
Lymphatic  glands 
Spleen     .... 
Thymus       .... 


PRACTICAL    WORK. 

PAGE 
I 

5 
6 


13 
14 

20 

30 
36 

42 

47 
52 
S3 
60 
62 
64 
65 
65 
67 

71 

75 
76 

77 
78 


PAGE 

Ductless  glands 

.     82 

Supra-renal  capsules         .... 

82 

Kidneys 

•      H 

Ureter 

.       SB 

Bladder 

.       89 

Skin 

.       90 

Spinal  cord  and  medulla  oblongata 

.       96 

Cerebellum  and  cerebrum 

.     100 

Nerve-ganglia 

.     102 

Eye 

.     104 

Cornea 

.     104 

Nose 

.     Ill 

Semicircular  canals  and  cochlea  . 

113 

Testis      ....... 

116 

Epididymis           ..... 

118 

Ovary      ....... 

120 

Fallopian  tube      ..... 

121 

Uterus     ...                 ... 

T2I 

Mammary  inland            .... 

122 

Milk        .    ' 

122 

Placenta       ...... 

122 

Deciduse          ...... 

124 

.Amnion       ....... 

124 

Umbilical  cord         ..... 

124 

INDEX 


125 


ABBREVIATIONS   USED  IN  THE    TEXT. 

(H)  means  /lig/i  power,  i.e.  a  magnifying  power  of  300  diameters  linear.  Such  a  magnifying  power  is 
obtained  by  using  a  No.  7  objective,  and  a  No.  3  eyepiece  of  Hartnack  with  the  tube  drawn  out  ;  or 
an  English  |-inch  objective  and  a  medium  eyepiece. 

(L)  means  l(nv  power,  i.e.  a  magnifying  power  of  65  diameters  linear.  Such  a  magnifying  power  is 
obtained  by  using  a  No.  3  objective  and  a  No.  3  eyepiece  of  Hartnack  with  the  tube  drawn  out  ;  or  an 
English  one-inch  objective  and  a  medium  eyepiece. 

T.  S.  and  L.  S.  mean  transverse  and  longitudinal  section  respectively. 


EXPLANATION 


OF 


THE     OUTLINE     PLATES. 


Plate  I.    BLOOD. 

( To  face  /.  4. ) 

Fig. 

1.  Coloured  blood-corpuscle  of  a  newt  or  frog  on  the  flat. 

2.  Coloured  blood-corpuscle  of  a  newt  or  frog  on  the  edge. 

3.  Effect  of  acetic  acid  on  a  newt's  corpuscle. 

4.  Effect  of  syrup  on  a  newt's  corpuscle. 

5.  Effect  of  tannic  acid  on  a  newt's  corpuscle. 

6.  Effect  of  water  on  a  newt's  corpuscle. 

7.  Effect  of  osmic  acid  and  picrocarmine  on  a  newt's  corpuscle. 

8.  Colourless  blood-corpuscles  of  a  newt  or  frog. 

9.  Human  blood-corpuscles  (coloured  and  colourless)  and  fibrin. 

10.  Human  coloured  blood-corpuscle  on  edge. 

1 1.  Human  coloured  blood-corpuscle  on  the  flat;  centre  in  focus  and  out  of  focus. 

12.  Human  coloured  blood-corpuscle  crenated. 

13.  Human  colourless  blood-corpuscles. 

14.  Effect  of  acetic  acid  on  these  corpuscles. 

15.  Effect  of  a  magenta  solution  on  these  corpuscles. 


Plate  II.    EPITHELIUM. 

(To  face  t>.  12.) 

1.  Squamous  epithelium  and  salivary  corpuscles  (mouth). 

2.  Squamous  epithelium  cuticle  of  newt  (intra-cellular  plexus  of  fibrils) 

3.  Silvered  epithelium  (mesentery  of  a  rabbit). 

4.  Columnar  epithelium  of  the  intestine. 

5.  Vertical  section  of  the  cornea. 

6.  Chalice  or  goblet  cell  from  the  small  intestine. 

7.  Columnar  epithelium  from  a  newt's  intestine. 

8.  Ciliated  epithelium  of  the  trachea. 

9.  The  free  ends  of  the  cells  of  fig.  8. 

10.  Bars  of  the  gills  of  a  mussel. 

11.  Transitional  epithelium. 

12.  Secretory  epithelium. 


Xll 


EXPLANATION  OF  PLATES. 


Fill. 


J- 

4- 

5- 
6. 


Plate  III.     CARTILAGE. 

{Tof.uep.  .6.) 

Hyaline  cartilage. 

Transverse  section  of  a  rib  of  a  small  mammal. 

White  fibro-cartilage  (intervertebral  disc.) 

Transverse  section  of  a  rabbit's  or  cat's  ear  (L). 

Transverse  section  of  a  rabbit's  or  cat's  ear  (H). 

Vertical  section  of  articular  cartilage  and  subjacent  bone. 


Plate  IV.     CONNECTIVE    TISSUE  AND    TENDON. 


(To  face  p.  24.) 

Fibres  of  elastic  tissue  (Lig.  nuchse  of  ox). 
Transverse  sections  of  these  fibres. 
Fresh  areolar  tissue. 
Effect  of  acetic  acid  on  fig.  3. 
Longitudinal  section  of  a  tendon  (H). 
Transverse  section  of  a  tendon  (L). 
Transverse  section  of  a  tendon  (H). 
Tendon  of  the  tail  of  a  rat  (L). 
Tendon-cells  from  the  tail  of  a  rat  (H). 


Plate  V.     OMENTUM  AND  FAT 

{To  face  p.  28.) 

1.  Omentum  of  a  rabbit  silvered. 

2.  Omentum  of  a  cat  silvered. 

3.  Vertical  section  of  foetal  skin  (developing  fat-cells  (L)). 

4.  Fat-cells  (H). 

5.  Fat-cells  after  the  action  of  ether. 

6.  Mucous  tissue. 

7.  Adenoid  tissue. 

Plate  VI.     BONE. 

[To  face  p.  32.) 

I  T.  S.  of  the  shaft  of  a  decalcified  bone  (L). 

2.  Part  of  fig.  I  with  (H). 

3.  L.  S.  of  a  bone  with  its  blood-vessels  injected. 

4.  T.  S.  of  dense  dry  bone. 

5.  Lacuna  of  dense  dry  bone  on  the  flat 

6.  T.  S.  of  the  shaft  of  a  foetal  bone. 

7.  V.  S.  through  the  line  of  ossification. 


Plate  VII.    MUSCLE. 

(To  face  p.  40.) 

1.  Non-striped  muscle  of  the  small  intestine. 

2.  Isolated  non-striped  muscle-cells. 

3.  Transverse  section  of  non-striped  muscle. 

4.  Cement-substance  of  non-striped  muscle,  silvered, 

5.  Non-striped  muscle-cells  from  the  mesentery  of  a  newt. 


EXPLANATION  OF  PLATES.  xiii 

Fio. 

6.  Sarcolemma  of  striped  muscle  (effect  of  water). 

7.  Effect  of  acetic  acid  on  striped  muscle  (frog). 

8.  Isolated  striped  muscular  fibre. 

9.  Muscle-discs. 

10.  Muscular  fibre  splitting  into  fibrils. 

1 1.  Striped  muscle  of  a  crab. 

1 2.  T.  S.  of  a  striped  muscle. 

13.  Blood-vessels  of  an  injected  muscle. 

Plate  VIII.    NERVE-FIBRES. 

(Tofacep.  iA-) 

1.  Fresh  meduUated  nerve-fibres  (myeline  drops). 

2.  Fresh  medullated  nerve-fibres  (nodes  of  Ranvier). 

3.  Effect  of  osmic  acid  on  nerve-fibres  (H). 

4.  The  incisures  of  Schmidt. 

5.  Effect  of  osmic  acid  (L). 

6.  Non-medullated  nerve-fibres. 

7.  Nerve-fibre  (medullated)  after  chromic  acid. 

8.  Small  nerve  silvered  (Ranvier's  crosses)  (L). 

9.  Nerve-fibre  silvered  (Ranvier's  crosses)  (H). 

10.  T.  S.  of  a  sciatic  nerve  (L). 

11.  T.  S.  of  a  sciatic  nerve  (H). 

12.  Concentric  lines  in  the  myeline  of  nerve-fibres. 

Plate  IX.    HEART  AND  BLOOD-VESSELS. 


(To  face  p.  SO.) 

Muscular  fibres  from  the  heart  of  a  mammal. 

T.  S.  of  the  muscular  fibres  of  the  heart  of  a  mammal. 

Muscle-cells  from  Purkinje's  fibres  of  a  sheep's  heart. 

Capillaries  silvered. 

Small  artery  silvered. 

Non-striped  muscle  in  a  small  artery. 

Small  artery  after  acetic  acid,  or  stained. 

V.  S.  of  a  middle-sized  artery. 

Plate  X.     TRACHEA  AND  LUNGS. 

(To  face  p.  56.) 
T.  S.  of  the  trachea  (L). 
Part  of  Fig.  i  with  (H). 
T.  S.  of  a  bronchus  and  blood-vessels  at  the  root  of  a  lung. 

Plate  XI.    LUNG. 

(To  face  p.  58.) 

Part  of  a  bronchus  (H). 
V.  S.  of  the  pleura  and  air-vesicles.  (L). 
Air- vesicles  of  a  lung  stained. 
Air-vesicles  of  a  lung  silvered. 
Injected  blood-vessels  of  a  lung  (L). 
V.  S.  of  a  foetal  lung. 


XIV 


EXPLANATION  OF  PLATES. 


Fia 
I 

2 

3 
4 


Plate  XII.     TONGUE  AND  TOOTH. 

(To  face  p.  62.) 

T.  S.  of  a  tongue — general  view  (L). 
V.  S.  of  a  tongue  with  papilla;  (H). 
Taste-bulbs  in  the  papillaj  foliatEE  (L). 
Taste-bulbs  (H). 
V.  S.  of  a  decalcified  tooth  and  jaw. 


Pl.\te  XIII.     SALIVARY  GLANDS  AND   STOMACH. 

(To  face  p.  66.) 

Sub-maxillary  gland  (mucous)  of  a  dog. 
V.  S.  cardiac  end  ot  the  stomach  (cat)  (L). 

3.  V.  S.  pyloric  end  of  the  stomach  (cat)  (L). 

4.  Peptic  gland  (H). 
T.  S.  of  the  peptic  glands  (H). 


Plate  XIV.     SMALL  AND  LARGE  INTESTINE. 

(To face f.  70.) 

V.  S.  of  the  small  intestine  (cat)  (L). 

V.  S.  of  an  intestinal  villus  (H). 

v.  S.  of  a  Lieberkiihn's  follicle. 

T.  S.  of  a  Lieberkiihn's  follicle. 

V.  S.  of  the  vermiform  appendix  (rabbit). 

V.  S.  of  the  mucous  membrane  large  intestine  (L). 


I. 
2. 
3- 

4- 

5- 
6. 


Plate  XV.     LIVER. 

(To  face  p.  72.) 

T.  S.  liver  and  its  capsule  (pig)  (L). 
T.  S.  liver  and  its  capsule  (rabbit)  (L). 
Columns  of  liver-cells  (H). 
Columns  of  liver-cells  (newt)  (H). 


Plate  XVI.    LIVER  AND  PANCREAS. 

(To  face  p.  74.) 

1.  Blood-vessels  of  a  liver  (rabbit)  injected  (L). 

2.  T.  S.  of  a  portal  canal  and  Glisson's  capsule. 

3.  Injected  bile-ducts. 

4.  Alveoli  of  pancreas. 


Plate  XVII.     LYMPHATICS. 

(To  face  p.  76.) 

Silvered  lymphatics  (small  intestine). 

Stomata  from  the  wall  of  a  frog's  great  lymph-sac. 

V.  S.  of  a  lymphatic  gland  (L). 

V.  S.  of  the  cortex  of  a  lymphatic  gland  (H). 

V.  S.  of  the  medulla  of  a  lymphatic  gland  (H). 

Adenoid  tissue  (H). 


EXPLANATION  OF  PLATES. 


XV 


Plate  XVIII. 


Fig. 
I. 

2. 

3- 
4- 


SPLEEN  AND  THYROID  GLAND. 

(To  face  p.  8o.) 


T.  S.  of  a  spleen  (cat)  (L). 

T.  S.  of  a  spleen  (H). 

T.  S.  of  a  Malpighian  corpuscle  (H). 

T.  S.  of  the  thyroid  gland. 


Plate  XIX.    KIDNEY. 

( To  face  A  86. ) 

1.  L.  S.  of  a  kidney  (L). 

2.  V.  S.  of  the  cortex  of  a  kidney  (L). 

3.  Bowman's  capsule  and  glomerulus  (H). 

Plate  XX.    KIDNEY  AND  SUPRA-RENAL  CAPSULE. 

(To  face  p.  88.) 

1.  V.  S.  of  the  cortex  (straight  and  convoluted  tubules). 

2.  Straight  tube  of  the  medulla. 

3.  Injected  blood-vessels  of  a  kidney. 

4.  V.  S.  of  a  supra-renal  capsule  (L). 

5.  V.  S.  of  a  supra-renal  capsule  (H). 


Plate  XXI.     SKIN. 

(To  face  p.  92.) 

1.  V.  S.  of  the  skin,  uninjected  and  injected  (L). 

2.  V.  S.  of  the  skin  (H). 

3.  Sweat-glands. 

4.  T.  S.  of  a  Pacini's  corpuscle. 


Plate  XXII.     SKIN  AND  HAIR. 

(To  face  p.  94.) 

V.  .S.  of  the  scalp  with  a  hair-follicle  (L). 
A  sebaceous  gland  (H). 
A  hair  without  medulla. 
A  hair  with  medulla. 


Plate  XXIII.     SPINAL  CORD. 

(To face  p.  98.) 

1.  T.  S.  of  the  spinal  cord. 

2.  Anterior  horn  of  the  spinal  cord  (H). 

3.  T.  S.  of  the  white  matter  of  the  cord  (H). 

Plate  XXIV.    BRAIN  AND  NERVE-GANGLIA. 

(To  face  p.  102.) 
V.  S.  of  the  cerebellum  (L). 
V.  S.  of  the  cerebellum  (H). 

3.  V.  S.  of  the  cerebrum  (L). 

4.  Nerve-cell  of  the  cerebrum  (H). 

5.  L.  S.  of  the  Gasserian  ganglion  (L). 

6.  Nerve-cell  of  fig.  5  without  its  capsule  (H). 
Nerve  cell  of  fig.  5  with  its  capsule  (H). 


xvi  EXPLANATION  OF  PLATES. 


Fig 
I 

2 

3 
4 
5 
6 

7 


Plate  XXV.     CORNEA  AND  CHOROID. 

{To  face  p.  I06.) 

V.  S.  of  the  cornea. 

Corneal  corpuscles  stained  with  gold  chloride. 

Cell-spaces  in  cornea  after  silver  nitrate. 

Lens-fibres  (man  or  dog). 

Lens-fibres  (cod-fish). 

Branched  pigment-cells  of  the  choroid. 

Hexagonal  pigment-cells  of  the  retina. 


PLATE  XXVL     RETINA  AND  CILIARY  MUSCLE. 

{To  face  p.  no.) 

T.  V.  S  of  the  retina  (cat)  (L). 

2.  V.  S.  of  the  retina  (frog)  (H). 

3.  Cones  of  retina  of  a  codfish  (H). 

4.  Ciliary  muscle  (L). 

Plate  XXVII.    NOSE  AND  COCHLEA. 

(To  face  p.  114.) 

1.  V.  S.  of  the  nasal  mucous  membrane  (H). 

2.  Isolated  epithelial  cells  of  the  olfactory  region  (H). 

3.  T.  S.  of  a  turn  of  the  cochlea  (L). 

4.  Part  of  fig.  3  with  (H). 

Plate  XXVIII.     TESTIS,  OVARY. 

{To  face  p.  liS.) 

1.  T.  S.  testis  (L). 

2.  Part  of  a  seminal  tubule  with  developing  spermatozoa  (H). 

3.  T.  S.  of  the  epididymis  (L). 

4.  Part  of  a  tubule  of  the  epididymis  (H). 

5.  A  spermatozoon. 

6.  V.  S.  of  the  ovary  (L). 

Plate  XXIX.     OVARY,  FALLOPIAN  TUBE,  UTERUS. 

(To  face  p.  120.) 

1.  T.  S.  of  a  Graafian  follicle  with  an  ovum  (H). 

2.  V.  S.  of  an  ovary  (H). 

3.  T.  S.  of  a  uterus  (L). 

4.  T.  S.  and  V.  S.  of  the  uterine  glands  (H). 

5.  T.  S.  of  part  of  a  Fallopian  tube  (H). 

Plate  XXX.    MAMMA,  MILK,  PLACENTA. 

{Tofacep.  122.) 

1.  Alveoli  of  a  mammary  gland  (H). 

2.  Milk  fresh  and  after  acetic  acid  (H). 

3.  Placental  villus  (L). 

4.  Placental  villus  (H). 

5.  T.  S.  of  the  umbilical  cord  (L). 

6.  Part  of  fig.  5  with  (H). 


DIRECTIONS  FOR  FILLING  IN   THE  DETAILS  IN    THE 

OUTLINE  PLATES. 

As  a  rule  only  the  outlines  of  the  leading  features  in  the  sections  are  indicated. 

The  outlines  are  not  drawn  to  any  definite  scale,  but  where  practicable  the  figures  are  of 
the  actual  size  as  seen  under  a  linear  magnifying  power  of  three  hundred  diameters  ( x  300) 
or  (H),  such  a  relation  being  noted  on  one  side  of  the  plates  devoted  to  the  elementary  tissues. 

The  details  are  to  be  filled  in  by  the  student  in  pencil  (H  or  HB)  and  coloured,  the  latter 
being  done  with  coloured  pencils  or  water  colour. 

Sufficient  dexterity  in  the  manipulation  of  water  colour  is,  I  find,  readily  acquired  by  the 
student,  as  very  few  pigments  suffice. 

All  the  necessary  colours  are  contained  in  the  shilling  boxes  of  moist  colour  made  by 
Reeves  and  Sons. 

A  preliminary  ground  colour  or  light  wash  can  be  laid  on  in  most  cases  to  great 
advantage. 

Evenness  of  colouring  is  ensured  by  operating  with  a  full  brush  upon  the  paper  placed  at 
a  gentle  slope.  Commencing  at  the  top  of  the  figure,  carry  the  brush  from  side  to  side  with 
a  steady  even  stroke  until  the  whole  is  coloured.  Invariably  allow  one  colour  to  dry  before 
another  is  applied. 

It  is  sometimes  convenient  to  use  a  dilute  solution  of  picrocarmine  as  a  groundwash,  whilst 
the  nuclei  and  other  parts  stained  red  may  be  tinted  with  solution  of  carmine.  A  logwood 
tint  is  obtained  by  mixing  a  small  quantity  of  carmine  with  a  good  blue,  such  as  ultra- 
marine. 

The  leading  parts  to  be  filled  in  are  indicated  in  the  text. 


ILLUSTRATIONS     IN     TEXT 


o>»;o 


FIG. 
I. 


4- 

5- 
6. 

7- 


9- 

lO. 

II. 

12. 

13- 

14. 
15- 


End  of  a  mounted  needle 

Scissors  curved  on  the  flat 

Ranvier's  arrangement  for  holding 
Drop-bottles  with  Reagents 

Hartnack's  Microscope,  No.  III.  A. 

Swift's  College  Microscope     . 

Nosepiece  made  by  Swift 

Microscopic  Paraffine  Lamp  . 

Pocket  Sheet  -  metal  Gauge  for 
Measuring  Cover-glasses 

Cover-glass  Measurer  of  Zeiss 

Camera  Lucida  of  Chevalier  . 

Camera  Lucida  (Zeiss)  . 

Simple  Warm  Stage 

Warm  Stage  and  Apparatus  for 
maintaining  a  constant  tempera- 
ture under  the  Microscope  . 

Section  of  a  Gas-chamber 

Ranvier's  Photophore    . 


PAGE 

FIG 

xix 

16 

xix 

17- 

XX 

18. 

xxi 

19. 

xxi 

xxii 

20. 

xxiii 

21. 

XXV 

XXV 

22. 

xxvii 

23- 

xxvii 

24. 

xxviii 

25- 

xxviii 

26. 

xxix 

XXXV 

27. 

16.     Swift's    simple    Dissecting   Micro- 
scope ..... 

Mr.  A.  B.  Stirling's  Microtome 

Ranvier's  Hand  Microtome    . 

Dr.    Rutherford's    Freezing  Micro- 
tome ...... 

20.     Williams'  (Quecket  Club)  Freezing 
Microtome  .... 

The  Ether  Freezing  Microtome  of 
Dr.  Bevan  Lewis 

Modified  Turntable 

Cannulse  for  Injecting    . 

Ludwig's    Mercury    Pressure-appa- 
ratus ...... 

Simple    Mercurial    Injection-appa- 
ratus ...... 

Injection -apparatus     with    water- 
pressure      

Subcutaneous  Syringe  for  Interstitial 
Injections,  and  the  Cannula 


PAGE 
XXXV 

xxxvii 
xxxvii 


.xli 

xlix 

Hi 

liii 

liv 

Iv 


INTRODUCTION. 


■■^^^XKo 


HISTOLOGICAL  REQUISITES. 


The  student  of  Histology  requires  the  following  apparatus  : — 

1.  A  compOTmd  microscope,  capable  of  magnifying  from  fifty  to  three   hundred   diameters 
linear. 

2.  Two  short,  strong,  sharp-pointed  needles,  fixed  in  wooden  handles, 
at  least  five  inches  long,  leaving  about  half  an  inch  of  their  point  pro- 
jecting (fig.  i).  Each  student  ought  to  make  these  needles  for  himself 
Ordinary  sewing  needles  may  be  conveniently  fixed  in  crotchet-holders. 

3.  A  pair  of  steel  forceps,  which  must  not  be  too  broad  at  the 
points. 

4.  A  hollow  ground  razor,  for  making  sections  with  the  hand  (p. 
xxxvi).     The  razor  '  made  for  the  army '  by  John  Heifor  is  the  best. 

5.  A  pair  of  sharp-pointed  scissors.     It  is  advantageous  for  some  purposes  to  have  a  pair 
curved  on  the  flat  (fig.  2). 


Fig, 


«,  End  of  a 
Mounted  Needle. 
Natural  Size. 


Fig.  2.    Scissors  Curved  on  the  Flat.     Natural  Size. 


6.  Three  or  four  small  camel-hair  brushes — duck  size — for  putting  on  the  cement  used  to 
seal  up  the  preparations.  It  is  convenient  to  keep  Nos.  2,  3,  4,  5,  and  6  in  a  small  box  or  in  a 
small  drawer  in  one's  vvork-table. 

7.  Glass  slides. — It  is  desirable  to  have  two  sizes.  The  glass  should  be  free  from  specks 
and  not  too  thick,  and  the  edges  ought  to  be  ground. 

{a)  Six  dozen  ground-glass  slides  3x1  inch. 

{b)  Three  „  „  3  x  I2  » 

S.  Cover-glasses. — I  prefer  the  circular  form,  though  many  observers  use  squares.  The 
one  point  of  importance  about  them  is  that  they  cannot  be  too  thin.  It  is  well  to  measure 
their  thickness,  and  this  can  be  easily  done  as  described  at  p.  xxv.  Messrs  Chance,  of 
Birmingham,   make   cover-glasses  of  three  thicknesses,  which  are  numbered  Nos.  i,  2,  and  3. 

b2 


XX 


INTRODUCTION. 


No.  I  is  the  thinnest,  and  is  what  ought  to  be  used.  They  vary  in  thickness  from  -004  to  008 
inch  in  thickness. 

(a)  \  oz.  extra  thin  f  inch  circles,  or  No.  i. 

{b)  \  oz.  extra  thin  i  inch  circles,  or  No.  i. 

{c)    \  oz.  extra  thin  \\  inch  circles,  or  No.  i. 

These  covers  ought  to  be  arranged,  according  to  their  size  and  thickness,  in  a  small  box 
divided  into  compartments  by  means  of  small  pieces  of  pasteboard.  This  the  student  can 
easily  make  for  himself. 

9.  A  pine-wood  box,  or  cabinet,  provided  with  trays,  so  that  the  specimens  may  lie  on  the 
flat.  The  trays  ought  to  be  made  both  for  large  and  small  slides,  and  the  cabinet  ought  to 
contain  at  least  si.x  dozen  slides.  Each  student  in  the  class  of  practical  physiology  mounts 
over  one  hundred  slides  in  a  session. 

10.  Reagents. — Each  student  ought  to  be  provided  with  a  small  wooden  framework  for 
holding  bottles,  containing  the  solutions  in  common  use. 

It  is  not  of  much  importance  what  kind  of  bottle  is  employed.  One-ounce  bottles  with 
moderately  wide  mouths,  and  fitted  with  a  glass  rod  passing  through  the  cork,  answer  ad- 
mirably.    The  solutions  most  commonly  employed  are  : — 

1.  \ pey  cent,  salt  solution  (p.  xxx). 

2.  Dilute  Acetic  Acid,  i  part  to  10  of  water. 

3.  Solution  of  Picrocarniinc  (p.  xliii). 

4.  Solution  of  ordinary  Carmine  (p.  .xliii). 

5.  Solution  of  Logwood  {t^.  xlii). 

6.  Clove  oil. — It  is  advantageous  to  use  a  camel-hair 
or  sable  brush  fi.xed  to  the  end  of  a  piece  of  wood  thrust 
through  the  cork. 

7.  Dammar  Mounting  Fluid  (p.  xlviii). 

8.  Glycerine. 

9.  Farranfs  Solution  (p.  xlviii). 

The  other  solutions  are  supplied  as  they  are  required. 
Ranvier    uses    a    very  convenient    arrangement    for 

holding  the  reagent  bottles  (fig.  3).  It  consists  of  a 
circular  glass  vessel,  about  four  inches  in  diameter  and 
two  inches  deep.  Into  its  mouth  a  perforated  plate  of 
cork  is  fitted,  and  in  this  cork  small  '  drop  bottles  '  are 
fixed,  and  the  whole  is  covered  with  a  glass  shade  to  keep  out  dust.  Anyone  can  easily  make 
such  an  arrangement  for  himself. 

10.  Narrow  strips  of  blotting  paper,  which  ougKt  to  lie  in  a  small  tray  attached  to  the 
stand  containing  the  reagents.  They  are  of  use  to  soak  up  surplus  fluid,  and  also  serve 
as  a  white  background,  when  such  is  required,  as  in  teasing  certain  coloured  preparations. 

Watch-glasses  and  small  glass  capsules  are  required,  and  the  student  will  find  the  instru- 
ments of  his  dissecting  case  very  useful.  Glass  pipettes  are  occasionally  required  to  remove 
fluid.  They  are  easily  made  by  heating  a  narrow  glass  tube  in  a  gas  flame,  and  then  drawing 
it  out. 


Fig.  3.  Ranvier's  Arrangement  for 
Holding  Drop-Bottles  with  Re- 
agents. 


INTRODUCTION. 


XXI 


ON    THE    USE    OF    THE    MICROSCOPE. 


GENERAL    DIRECTIONS. 


The  microscope  used  in  the  Physiological  Laboratory  of  the  University  of  Aberdeen  is 
Hartnack's,  model  No.  III.  A.  (fig.  4)  with  an  eyepiece,  or  ocular,  No.  3,  and  two  objectives,  or 
lenses,  Nos.  3  and  7.  The  combination  of  ocular  No.  3  with  objective  No.  3,  gives  a  mag- 
nifying power  of  about  65  diameters  linear,  and  ocular  No.  3  and  objective  No.  7  about  300 
diameters,  with  the  tube  drawn  out.  These  powers  are  quite  sufficient  for  beginners,  and, 
indeed,  for  most  of  the  ordinary  work.  This  microscope  costs  about  7/.  Higher  magnifying 
powers  aie  supplied  when  they  are  required. 


Fig.  4.    Hartnack's  Microscope  No.  III.  A. 
One-fiflli  Natural  .Size.     The  Condenser  is  attached 
to  the  Body  of  the  Microscope. 


Fig.  5.    .Swift's  College  Microscope. 
With  a  Condenser  attached  to  the  Stage  for  examining 
opaque  objects. 


The  magnifying  power  can  be  diminished  or  increased  by  pushing  or  pulling  out  the  draw- 
tube.     In  fig.  4,  the  draw-tube  is  represented  as  drawn  out. 

In  Hartnack's  microscope,  the  stand  has  a  horse-shoe  form,  but  many  prefer  the  tripod  form 
(fig.  5)  as  made  by  Messrs.  Swift  and  Son,  University  Street,  London.  In  this  microscope  the 
tube  is  cloth-lined,  so  that  it  moves  easily,  and  the  stage  is  covered  with  a  glass  plate  which 
enables  the  slide  to  move  easily  on  it,  and  prevents  it  from  being  acted  on  by  acids. 

The  object-glass,  objective,  or  lens,  is  the  most  important  part  of  a  microscope,  and  it  is 
necessary  to  see  that  a  good  lens  be  procured.  The  lens  should  be  tested  by  a  competent 
microscopist  before  it  is   purchased.     Coloured  blood-corpuscles  and   salivary  corpuscles  are 


xxii  INTRODUCTION. 

better  '  test  objects'  than  the  markings  of  diatoms.  It  is  most  important  to  have  a  good  high 
power  such  as  Hartnack's  No  7,  equal  to  about  an  English  \  inch.  If  a  higher  power  be 
required,  Zeiss'  E  =  g  inch,  and  his  F  =  j\7  inch,  are  excellent  lenses. 


IMMERSION    LENSES. 

Ordinary  lenses  are  '  dry,'  that  is,  air  is  the  medium  between  the  object  and  the  objective. 
For  higher  powers,  lenses  are  specially  constructed  so  that  a  fluid  medium  is  placed  between  the 
object  and  the  objective.  The  effect  of  this  is  to  collect  a  larger  number  of  rays  of  light 
passing  through  the  object,  and  so  increase  the  illumination  of  the  field  of  the  microscope. 
Water  was  formerly  employed,  and  lenses  were  specially  constructed  for  use  with  water 
as  the  medium.  Recently,  however,  oil  immersion  lenses  have  been  constructed  by  Zeiss  of 
Jena,  and  his  Jy  inch  is  the  best  lens.  These  lenses  are  rapidly  taking  the  place  of  water 
immersion  lenses.  Similar  lenses  are  made  by  Messrs  Powell  and  Lealand.  Cedar  oil  is  the 
medium  employed  for  these  lenses.  A  drop  of  oil  is  placed  on  the  lens,  and  the  tube  is  then 
lowered  till  the  oil  comes  in  contact  with  the  slide,  and  the  object  is  focussed  in  the  ordinary 
way.  They  increase  the  sharpness  and  brilliancy  of  objects  immensely.  As  the  cedar  oil 
dissolves  dammar,  all  preparations  to  be  examined  with  oil  lenses  must  be  sealed  up  with 
marine  glue,  which  is  not  acted  on  by  cedar  oil.  It  is  not  necessary  for  the  student  to  get 
these  expensive  lenses  ;  they  are  supplied  when  they  are  required. 

A  nosepiece  is  a  simple  device  (fig.  6)  by  means  of  which  the  unscrewing  of  one  objective 
to  use  another  is  obviated.     Either  the  high  or  the  low  power  can  be  u.sed.     It  is  necessary 

that  both  lenses  be  accurately  centred.  The  nosepiece  is 
screwed  to  the  tube  of  the  microscope,  and  to  it  the  lenses  are 
screwed.  A  nosepiece  may  likewise  be  made  to  carry  three 
or  four  lenses. 

Eyepieces. — Nos.  3  and  4  are  supplied  with  Hartnack's 
microscope  but  it  is  well  always  to  use  the  deep  eyepiece, 
or  No.  3,  i.e.  the  less  powerful  of  the  two. 

THE   ADJUSTMENTS    OF    A   MICROSCOPE. 

In  every  microscope  there  are  two  adjustments  : — 
{a)  The  coarse  adjustment,  which  serves  to  bring  the 
Fig.  6.  Nosepiece  made  by  Swift,  ^^ject  roughly  within  focal  distance.  In  Hartnack's  micro- 
scope, this  is  accomplished  by  rotating — not  pushing — one 
tube  inside  another  with  the  hand.  This  is  b)'  far  the  most  convenient  method.  As  the 
tube  of  this  microscope  is  not  cloth-lined,  the  two  surfaces  ought  to  be  kept  perfectly  clean  to 
ensure  easy  motion.  If  the  tube  does  not  move  easily  enough,  rub  it  with  a  little  almond 
or  watchmaker's  oil,  and  move  it  upwards  and  downwards  till  it  glides  easily.  Wipe  off  the 
surplus  oil  with  a  cloth. 

{b)  The  pine  adjustment,  is  accomplished  by  rotating  a  milled  head  or  screw  placed  at  the 
upper  part  of  the  pedestal  of  the  microscope.  It  serves  to  bring  the  object  accurately  into 
focus  after  its  outline  has  been  brought  into  view  by  means  of  the  coarse  adjustment. 

The  stage,  on  which  the  object  is  placed,  ought  to  be  fixed,  and  at  least  three  inches  in 
breadth.  In  Hartnack's  microscope  it  is  placed  at  a  very  convenient  height  above  the  table, 
so  that  when  the  outer  surface  of  the  left  hand  is  resting  on  the  table,  the  thumb  and  fore- 


INTRODUCTION. 


xxm 


graduated   apertures,  are 


finger  can  be  used  to  move  the  slide  on  the  stage.     In  this  way  all  unnecessary  fatigue  of  the 
arm  is  avoided.     In  Swift's  microscope  it  is  glass-plated. 

The  diaphragm  is  placed  under  the  stage,  and  consists  of  a  circular  plate,  with  a  graduated 
series  of  round  apertures  in  it.  When  using  a  high  power,  turn  round  the  diaphragm  until 
a  small  aperture — usually  the  second  smallest — is  directly  under  the  hole  in  the  centre  of  the 
stage.  A  small  catch  indicates  when  this  is  accomplished.  When  using  a  low  power,  employ 
a  large  aperture  of  the  diaphragm. 

On  the  more  expensive  microscopes  small  stops,  or  tubes  with 
supplied  with  the  lenses.     They  are  slipped  into  a  hole  in  the  stage 
from  below,  but  care  must  be  taken  that  they  are  accurately  centred, 
so  that  their  apertures  are  exactly  under  the  objective. 

The  illumination.  This  is  accomplished  by  means  of  a  mirror 
placed  under  the  stage.  Never  employ  direct  sun-light ;  rather 
employ  light  from  a  white  cloud.  In  selecting  a  site  to  work  with 
the  microscope,  try  to  secure  a  north  light. 


ARTIFICI.A.L    ILLUMINATION. 

Under  certain  circumstances  it  is  necessary  to  use  a  lamp 
which  burns  either  gas  or  paraffine.  I  find  that  an  ordinary  gas 
jet,  fitted  with  a  Sugg's  burner,  surrounded  by  a  chimney  of 
pale  blue  glass,  answers  admirably.  Figure  7  shows  a  form  of 
microscopic  lamp  which  burns  paraffine.  It  is  also  very  convenient. 
The 
funnel.     It  is  made  by  Swift, 


ight  is  better  when  it  is  transmitted  through  a  light  blue  glass 


An  inexpensive  convenient  light  is  obtained  from  an  ordinary  Fig.  7.  Microscopic  Paraffine 
paraffine  lamp  with  a  flat  wick.  '      (  «'i  )■ 

CLEANING    OF    THE    LENSES    OR   OBJECTIVES. 

The  lenses  are  optically  the  most  important  part  of  a  microscope.  Never  unscrew  the 
separate  parts  of  the  objective  or  lens.  If  there  be  any  dimness,  it  is  sure  to  be  due  to  dirt 
on  the  outer  lenses.  Carefully  rub  the  lens  with  a  piece  of  chamois  leather,  which  should  be 
kept  ready  at  hand.  A  convenient  way  is  to  tie  it  to  the  stand  of  the  microscope.  If  a  drop 
of  dammar  chance  to  be  on  the  lens,  it  must  be  cautiously  removed  by  applying  a  drop  of 
clove  oil  and  removing  it  quickly  with  chamois  leather.  It  is  necessary  to  remove  the  clove 
oil  at  once,  as  the  glasses  which  compose  the  lens  are  soldered  together  with  Canada  balsam 
which  would  be  dissolved  by  the  clove  oil.  A  dilute  solution  of  ammonia  applied  to  the  lens 
with  chamois  leather  or  cotton  wool  is  an  excellent  method  of  removing  the  greasy  film  which 
sometimes  coats  glass.  .  The  surface  of  a  freshly-fractured  piece  of  elder  pith  answers  the 
same  purpose  admirably. 


METHOD    OF    FOCUSSING   AN    OBJECT. 

It  is  extremely  desirable  for  the  student  of  histology  to  begin  at  once  by  learning  to  keep 
botli  eyes  open  when  examining  an  object.  After  putting  on  the  proper  lens,  arrange  the 
mirror  under  the  stage  so  as  to  direct  a  beam  of  light  up  the  tube  of  the  microscope  and 
illuminate  the  field.     See  that  the  field  is  brightly  illuminated,  and  that  no  specks  are  present. 


xxiv  INTRODUCTION. 

If  specks  are  present,  it  may  be  easily  determined  as  to  whether  they  are  on  the  ocular  (eye- 
piece) or  on  the  objective  ;  if  they  arc  on  the  eyepiece,  they  will  necessarily  move  when  it  is 
moved.     They  must  be  removed  by  gentle  cleaning  with  chamois  leather. 

(rt)  Low  Power  (L).— With  the  No.  3  objective.  Place  the  tube  of  the  microscope  so  that 
the  lens  is  at  least  an  inch  and  a  half  above  the  object.  Keep  looking  through  the  micro- 
scope, and,  whilst  doing  so,  keep  the  slide  on  the  stage  in  motion  with  the  left  hand,  and 
rotate  or  twist  the  tube  downwards — always  from  right  to  left — until  the  outline  of  the  object 
comes  distinctly  into  view.     Then  employ  the  fine  adjustment,  and  accurately  focus  the  object. 

(1^)  High  Power  (H). — With  the  No.  7  objective.  Place  the  lens  a  quarter  of  an  inch  above 
the  object,  and  then  slowly  rotate  the  tube  downwards,  keeping  the  object  on  the  stage  in 
motion  all  the  time  with  the  thumb  and  fore-finger  of  the  left  hand,  until  the  object  just  comes 
into  view,  then  employ  the  fine  adjustment.  As  already  indicated  the  magnifying  power  may 
be  increased  by  pulling  out  the  draw-tube. 

Movement  of  the  Slide  on  the  Stage. — This  is  done  most  conveniently  by  means  of  the 
thumb  and  fore-finger  of  the  left  hand.  Never  employ  two  hands  to  move  a  slide  ;  it  is  quite 
unnecessary.  By  practice  wonderful  precision  of  movement  is  acquired.  Two  small  clips  are 
placed  on  the  stage,  which  serve  to  fix  the  slide,  so  that  it  remains  in  any  position.  All  the 
cumbrous  mechanical  appliances  formerly  in  fashion  for  moving  the  slide  on  the  stage  are  quite 
unnecessary. 

GENERAL   RULES    FOR   THE    EXAMINATION    OF   AN    OBJECT. 

Always  begin  the  examination  of  an  object  with  a  low  power,  in  order  to  obtain  a  general 
survey  of  the  arrangement  of  the  parts.  If  it  be  desired  to  study  any  particular  part,  place  it 
directly  in  the  centre  of  the  field  of  the  microscope  by  means  of  the  low  power,  and  then  ex- 
amine it  with  a  higher  power,  taking  care  in  the  latter  instance  to  use  a  small  aperture  of  the 
diaphragm. 

PREPARATION    OF   A   SIMPLE    OBJECT. 

A  drop  of  blood  or  milk  serves  the  purpose  very  well.  Clean  a  slide  carefully.  Dip  the 
slide  in  water,  and  rub  both  surfaces  at  once  with  a  towel  which  has  been  washed  several 
times,  so  as  to  get  rid  of  all  adherent  threads.  It  is  often  convenient  for  the  beginner  to  use 
a  coloured  towel — say,  a  red  one — for  if  he  finds  a  coloured  thread  in  the  field  of  the  micro- 
scope, he  knows  that  it  has  come  from  the  towel.  Place  a  small  drop  of  the  fluid  to  be  ex- 
amined on  the  centre  of  a  slide  by  means  of  a  clean  glass  rod.  Apply  a  clean  cover-glass, 
taking  care  to  prevent  the  entrance  of  air-bubbles. 


MEHODS    OF   APPLYING   A   COVER-GLASS. 

Seize  by  the  edge  a  previously  cleaned  cover-glass  with  forceps,  and  allow  the  drop 
of  fluid  to  come  in  contact  with  the  edge  of  the  cover  farthest  removed  from  the  forceps  ; 
gradually  and  slowly  allow  the  cover-glass  to  descend  obliquely  until  the  fluid  forms  a  uniform 
stratum  under  the  cover.  There  ought  to  be  just  a  sufficient  amount  of  fluid  to  fill  out  the 
space  between  the  slide  and  cover-glass.  When  forceps  are  not  at  hand,  the  cover  may  be 
applied  with  the  fingers.  Take  a  cover  by  the  margins,  rest  one  edge  on  the  slide  close  to  the 
drop,  allow  the  fluid  to  touch  the  under  surface  of  the  edge  of  the  cover,  and  slowly  permit 
the  cover  to  descend  obliquely. 


INTRODUCTION. 


XXV 


METHOD    OF    CLEANING    COVER-GLASSES. 

Not  unfrequently  covers  are  coated  with  a  dull  film,  which  obscures  the  passage  of  the 
light.  To  remove  this  film  place  the  cover-glasses  for  an  hour  or  two  in  a  beaker  containing 
strong  sulphuric  or  nitric  acid.  Pour  off  the  acid,  and  wash  the  glasses  thoroughly  with 
water  until  all  the  acid  is  removed.  They  may  then  be  placed  in  methylated  spirit,  and  as 
each  cover-glass  is  taken  out  it  is  cleaned  with  chamois  leather  or  a  well-washed  silk  hand- 
kerchief This  is  accomplished  by  rubbing  the  surfaces  of  the  cover-glass  with  the  hand- 
kerchief, held  between  the  thumb  and  forefinger  of  the  left  hand.  The  cover  ought  always  to 
be  lifted  by  the  edges,  and  never  allowed  to  lie  on  the  flat  after  it  is  cleaned,  as  particles  of  dirt 
are  apt  to  adhere  to  it.  Tilt  it  up  against  something  until  required.  For  inch-and-a-quarter 
cover-glasses  a  good  plan  is  to  have  two  perfectly  smooth  pieces  of  wood,  covered  with 
chamois  leather,  and  to  place  the  cover-glass  between  them,  and  rub  it  until  cleaned. 


ON    MEASURING   COVER-GLASSES. 

As  already  indicated.  No.  i  cover-glasses  do  very  well  for  the  ordinary  purposes  of  a 
student.  When  very  high  powers  are  used,  it  is  well  to  know  the  thickness  of  the  cover-glass 
employed.     Cover-glasses  may  be  rapidly  measured  in  two  ways. 

{a)  Micrometer  caliper. — This  little  American  instrument  (fig.  8)  is  sold  by  Messrs.  Chas. 
Churchill  and  Co.,  28,  Wilson  Street,  London.  It  is  used  for  measuring  the  thickness  of  fine 
brass  plates.  The  piece  in  the  form  of  the  letter 
U  has  a  projecting  hub,  a,  at  one  end.  Through 
the  two  ends  are  tapped  holes,  in  one  of  which 
is  the  adjusting  screw,  B,  and  in  the  other  the 
gauge  screw,  C.  Attached  to  the  screw,  C,  is 
a  thimble,  D,  which  fits  over  the  exterior  of  the 
hub,  a.     The  end  ofthis  thimble  is  bevelled,  and 


Fig.  S.    Pocket  Sheet-metal  Gauge  for  Measuring 
Cover-glasses.     Natural  Size. 


Fig.  9.   Cover-glass  Measurer  of  Zeiss. 


the  bevelled  edge  graduated  into  twenty-five  parts,  and  figured  O,  5,  10,  15,  20.  A  line  of 
graduation,  40  to  the  inch,  is  also  made  upon  the  outside  of  the  hub,  a,  the  line  of  these 
divisions  running  parallel  with  the  centre  of  the  screw,  C,  whilst  the  graduations  on  the 
thimble  are  circular.  The  pitch  of  the  screw,  C,  being  40  to  the  inch,  one  revolution  of  the 
thimble  opens  the  guage  i-40th  or  25-ioooths  of  an  inch.  The  divisions  on  the  thimble  are 
then  read  off  for  any  additional  part  of  a  revolution  of  the  thimble,  and  the  number  of  such 
divisions  are  added  to  the  turn  or  turns  already  made  by  the  thimble,  allowing  25-1000  for 
each  graduation  on  the  hub,  a. 

c 


xxvi  INTRODUCTION. 

Measure  an  ounce  of  No.  i  cover-glasses,  and  put  aside  all  those  under  "004  inch  for 
use  with  high  powers. 

{J})  The  cover-glass  measurer  (Deckglass-tastcr)  of  Zeiss  (fig.  9)  is  a  very  convenient  appa- 
ratus, which  has  a  movable  disc,  d,  divided  into  degrees.  The  cover-glass,  c,  is  placed  between 
the  ends  of  two  screws,  and  the  disc  is  rotated  till  the  screw  just  holds  the  cover.  By  an 
ingenious  arrangement  of  teeth  the  handle  is  thrown  '  out  of  gear '  when  this  occurs.  It  gives 
the  thickness  of  a  cover-glass  in  parts  of  a  millimetre. 


HOW   TO    PLACE   A    SECTION    ON    A    SLIDE. 

The  section  ought  to  be  placed  in  a  bowl  of  water — preferably  a  white  bowl — as  it  admits 
of  the  section  being  easily  seen.  Take  a  slide,  and  plunge  it  for  three-quarters  of  its  length 
into  the  water,  and  with  a  needle  in  the  right  hand  gently  guide  the  section  on  to  the  slide, 
which  ought  to  be  held  in  a  slanting  direction.  The  section  is  thus  easily  _/?i3i7/'(:rt' on  to  the 
slide,  and  thus  all  unnecessary  tearing  of  it  is  avoided.  The  needle  is  used  to  hold  the  section 
on  the  slide  whilst  the  latter  is  being  removed  from  the  water.  Remove  the  surplus  water 
with  a  towel.  To  the  section  on  the  slide  the  necessary  staining  reagents  may  be  applied. 
Sections  may  easily  be  treated  with  alcohol  and  clove  oil  on  a  slide.  This  method  is 
infinitely  preferable  to  lifting  sections  from  clove  oil,  etc.,  even  with  the  most  approved  form 
of '  section-lifters.' 

LABELLING    OF    MICROSCOPIC    OBJECTS. 

This  is  most  important.  Every  preparation,  when  mounted,  ought  to  be  properly  labelled. 
Small  paper  labels  are  affixed  to  the  slide,  and  on  these  are  marked  the  nature  of  the  section, 
whether  it  is  a  transverse  (T.  S.)  or  a  longitudinal  section  (L.  S.)  ;  any  special  points  in  its  struc- 
ture ;  how  it  was  prepared  and  in  what  medium  it  is  mounted.  The  thickness  of  the  cover- 
glass  in  special  cases  ought  to  be  indicated.  By  using  a  series  of  symbols,  as  Pc  for 
picrocarmine,  Lg  for  logwood,  G  for  glycerine,  D  for  dammar,  and  F  for  Farrant's  solution, 
much  important  information  can  be  put  upon  one  small  piece  of  paper. 


ON    MAKING   DRAWINGS   OF    MICROSCOPIC 
PREPARATIONS. 

It  is  of  the  utmost  importance  that  each  student  should  make  a  sketch  of  many,  not  neces- 
sarily of  all,  the  preparations  he  mounts.  This  is  the  only  method  of  ensuring  that  he  sees 
what  is  to  be  seen.  The  outline  plates  are  designed  to  aid  the  student  in  this  task.  Once  the 
student  has  made  a  drawing  of  the  details  of  a  preparation  he  is  not  likely  to  forget  them.  It  is 
far  more  important  that  each  student  should  thoroughly  understand  a  selected  series  of  pre- 
parations than  that  he  should  carry  off  with  him  a  couple  of  hundred  slides,  few  of  which  he 
understands. 

Various  instruments  are  used  for  making  drawings  of  microscopic  preparations,  but  it  is  to 
be  remembered  that  they  are  in  many  cases  more  useful  for  giving  the  actual  size  of  an  object 
as  seen  with  a  certain  combination  of  lenses,  and  for  giving  the  general  outline  of  a  section, 
than  for  the  individual  details. 


INTRODUCTION. 


xxvu 


Camera  Lucida  of  Chevalier. — This  instrument  (fig.  lo)  is  made  by  Hartnack.     Instead  of 
the  eyepiece,  the  tube  s'  is  fixed  in  the  tube  of  the  microscope.     The  observer   looks  through 

the  small  prism,  and  sees  the  outline  of  the 
section  thrown  on  the  table,  or  on  a  piece  of 
paper  laid  on  the  table,  close  to  the  microscope. 
A  shade  is  placed  in  front  of  this,  and  must  be 
so  adjusted  that  too  much  light  does  not  fall 
on  the  paper.  The  contour  is  then  mapped 
out  with  a  pencil. 


Fig.   10.    Camera  Lucida  of  Chevalier. 
a  the  Instrument  ;  l>  a  section,  showing  the  prisms. 


Fig.   II.    Camera  Lucida  (Zeiss). 

The  ring  of  brass  fixes  it  to  the  tube  of  the  Microscope, 

and  the  aperture  a  is  placed  over  the  Eyepiece. 


Other  Cameras. — Much  simpler  forms  are  the  camera  lucida  of  Nachet,  or  the  one 
made  by  Zeiss  (fig.  1 1).  The  ocular  is  left  in  its  place,  and  the  camera  is  fixed  to  the  tube  of 
the  microscope  by  the  ring  of  brass.  The  aperture  a  is  placed  immediately  over  the  centre  of 
the  eyepiece.  On  looking  through  this  aperture  the  surface  of  the  table  in  front  of  the  micro- 
scope is  seen  at  the  same  time  as  the  object.  The  drawing-paper  is  placed  on  an  inclined 
plane  on  this  part  of  the  table,  and  the  outline  of  the  object  is  sketched,  whilst  the  details  are 
afterwards  filled  in  without  the  camera. 

Neutral  tint  reflector. — This  simply  consists  of  a  small  cap,  which  fits  on  the  end  of  the 
microscope,  and  in  it  is  placed  a  piece  of  glass,  at  an  angle  of  45°.  The  microscope  tube 
must  be  horizontal.  On  looking  through  the  glass  the  image  of  the  object  is  seen  on  the 
paper.  The  same  precautions  with  regard  to  the  modification  of  the  light  on  the  paper  must 
be  taken  as  for  Chevalier's  camera. 

It  is  not  even  necessary  to  have  a  brass  cap  to  fit  on  to  the  microscope  ;  an  ordinary  cover- 
glass,  fixed  to  the  eyepiece,  by  means  of  modeller's  wax,  at  an  angle  of  45°,  answers  the  purpose 
admirably. 

METHODS   OF   STUDYING   THE    PROPERTIES   OF 
LIVING   PROTOPLASM. 


THE   MOIST   CHAMBER. 


Moist  chamber  of  v.  Recklinghausen. — This  is  easily  made  by  cutting  off  the  lower  end  of 
an  ordinary  glass  cylinder,  such  as  is  used  for  a  paraffine  lamp.  Through  the  lower  part  of  the 
cylinder  the  tube  of  the  microscope  is  placed.     The  lower  end  of  the  cylinder  is  ground  flat, 


XXVIU 


IN  TROD  UC  TION. 


and  rests  on  a  plate  of  glass,  to  which  it  is  fitted  air-tight  with  oil.  The  inner  surface  of  the 
cylinder  is  partly  coated  with  filter  paper  moistened  with  water,  so  that  the  air  in  the  enclosed 
space  is  always  kept  saturated  with  watery  vapour.  To  close  the  space  all  that  is  required 
is  to  tie  a  layer  or  two  of  sheet  caoutchouc  over  the  lower  end  of  the  tube  of  the  microscope, 
and  the  upper  end  of  the  glass  cylinder.  The  object  to  be  examined  is  placed  on  the  floor 
of  this  chamber,  and  may  be  examined  either  with  or  without  a  cover-glass. 

Simple  moist  chamber. — A  very  simple  chamber  may  be  made  by  fixing  a  ring  of  glass,  a 
quarter  of  an  inch  in  height  and  seven-eighths  of  an  inch  in  diameter,  to  an  ordinary  slide  by 
means  of  dammar  solution.  A  drop  of  water  is  placed  in  the  chamber,  which  is  then  covered 
with  a  cover-glass,  on  which  the  fluid  to  be  examined  is  placed.  The  cover-glass  is  so  applied 
that  the  drop  of  fluid  hangs  into  the  chamber,  as  is  shown  in  fig.  14. 


METHODS   OF   APPLYING    HEAT   TO   MICROSCOPIC   OBJECTS. 

Various  forms  of  warm  stages  are  in  use  ;  Max  Schultze  was  the  first  to  use  them. 

Simple  form  of  zvarm  stage. — This  consists  of  a  thin  sheet  of  copper,  three  inches  long  and 
one  and  a  half  inches  broad  (fig.  12).  In  the  centre  is  a  hole  half  an  inch  in  diameter.  From 
one  side  of  the  plate  a  narrow  copper  arm,  five  inches  in  length,  projects.  This  is  placed  on 
the  stage  of  the  microscope,  and  on  it  is  placed  the  preparation  to  be  observed.  If  it  be  blood 
(p.  4),  the  cover-glass  must  be  sealed  round  with  oil  to  prevent  evaporation,  and  the  whole 
is  fixed  by  the  clips  on  the  microscope.     Heat  is  applied  to  the  brass  arm  by  means  of  a 


i 


^iH 


mm 


Fig.  12.    Simple  Warm  Stage. 
Half  Natural  Size. 


Fig.  13.    Warm  Stage  and  Apparatus  for  Maintaining  a  Constant 
Temperature  under  the  Microscope  (Schafer). 


spirit-lamp,  and,  in  order  that  it  may  not  be  over-heated,  a  piece  of  a  mixture  of  cocoa- 
butter  and  white  wax  is  placed  on  the  copper  plate  near  the  preparation.  This  mixture  is 
so  made  as  to  melt  at  a  temperature  of  30°  C.  When  it  melts  the  source  of  the  heat  must 
be  removed.  This  does  very  well  for  the  purposes  of  the  student ;  anyone  can  make  it  for 
himself  from  sheet  copper.  For  accurate  experiments  it  is  necessary  to  have  a  thermometer, 
so  that  the  degree  of  heat  can  be  accurately  measured.  Such  an  arrangement  is  found  in 
Strieker's  warm  stage. 


INTRODUCTION.  xxix 

Mr.  Schdfer's  constant  temperature  apparatus. — The  following  apparatus,  devised  by  Mr. 
Schafer,  is  an  excellent  one  (fig.  13).  It  consists  of  a  brass  box,  a,  just  like  a  Strieker's 
warm  stage,  having  a  hole  in  its  centre  to  admit  light  to  the  microscope.  In  it  is  a  ther- 
mometer, b.  This  box  lies  on  the  stage  of  the  microscope.  It  is  connected  by  elastic  tubes 
with  a  hollow  jacket,/,  and  the  whole  system  is  filled  with  water  previously  boiled  to  expel 
the  air.  The  water  is  warmed  at  ^  by  a  small  gas-flame,  the  size  of  which  is  regulated  by 
a  modified  Bunsen's  regulator,  (/.  By  means  of  the  screw  e  the  temperature  is  regulated  at 
starting. 


METHOD    OF   APPLYING   VAPOURS   AND    GASES. 

A  simple  apparatus  is  described  at  p.  11  for  applying  the  vapour  of  ether  or  chloroform 
to  an  object,  e.g.  to  cilia.  It  is  made  in  the  same  way  as  the  moist  chamber  described  at 
p.  xxviii. 

If  gases,  such  as  o.xygen  or  carbonic   acid,  are  to   be   applied,  two  tubes  require  to  open 


Fig.  14.    Section  of  a  Gas  Chamber,     a.  Entrance  ;  b,  Exit  tube  ;  c,  Object  ; 
d.   Cover-glass.     Natural  size. 

into  the  glass  cell,  one  for  the  entrance  of  the  gas,  and  the  other  for  its  exit.  Fig.  14  shows 
a  section  of  such  an  arrangement.  The  cover-glass,  with  the  object  on  it  is  inverted  over  the 
cell.     Strieker's  warm  stage  is  usually  so  made  that  it  can  be  used  for  this  purpose. 


XXX  INTRODUCTION. 


ON    THE   PREPARATION  OF   TISSUES  FOR 
MICROSCOPIC  EXAM  IN  A  TION. 


FRESH    TISSUES. 

Fresh  tissues  as  a  rule  are  unsuited  for  examination  with  the  microscope,  though  under 
certain  circumstances  it  is  advisable  to  examine  a  fresh  tissue.  When  this  is  done,  the  tissue 
must  be  examined  in  a  fluid  which  will  alter  its  characters  as  little  as  possible  ;  such  fluids 
are  called 


NORMAL   OR    INDIFFERENT    FLUIDS. 

They  closely  resemble  in  composition  some  of  the  fluids  in  which  the  tissues  of  the  body 
are  bathed. 

1.  Aqueous  humour  of  the  eye. — This  is  easily  obtained  by  puncturing  the  cornea  of  an  eye- 
ball removed  from  an  ox  newly  killed. 

2.  Blood-serum.— Pour  blood  into  a  tall  vessel,  and  allow  it  to  coagulate.  After  the  blood 
coagulates,  run  a  knife  between  the  upper  margin  of  the  clot  and  the  vessel,  to  permit  the  clot 
to  contract  and  sink  in  the  serum,  which  will  be  squeezed  out  of  the  clot.  After  twenty-four 
hours  draw  off  the  yellow-coloured  serum  with  a  pipette. 

3.  Iodised  serum. — Add  iodine  to  blood-serum,  prepared  as  above,  until  the  fluid  is  of  a 
distinctly  yellow  colour.  This  fluid  alters  the  tissues  slightly,  however,  and  colours  them 
yellow.     A  similar  solution  may  be  made  by  adding  iodine  to  amniotic  fluid. 

4.  Salt  solution  (three-quarter  per  cent.). — Dissolve  7-5  grammes  sodic  chloride  in  1,000 
c.c.  of  distilled  water.  This  is  by  far  the  most  convenient  fluid  to  employ.  Its  composition 
is  so  near  that  of  lymph — the  fluid  normally  bathing  the  tissues — that  it  alters  fresh  tissues 
very  slightly. 

HOW   TO    HARDEN    TISSUES. 

GENERAL   DIRECTIONS. 

Tissues,  however,  generally  require  to  be  hardened  before  they  can  be  cut  into  sections 
for  examination.     This  is  accomplished  by  exposing  the  tissue  to  the  action  of  certain  fluids. 

The  Material  to  be  employed. — The  tissues  should  in  all  cases  be  as  fresh  as  possible. 
When  possible  the  tissues  ought  to  be  obtained  from  the  human  body,  though  these  in  all 
cases  may  not  be  available.  It  is  therefore  necessary  to  use  the  corresponding  organs  of  some 
of  the  lower  animals,  e.g.,  cat,  dog,  rabbit,  or  pig.     Practically  their  structure  is  identical  with 


INTRODUCTION.  xxxi 

that  of  man.  The  tissue  to  be  hardened  ought  to  be  cut  into  small  pieces— half  to  one  inch 
square — by  means  of  a  sharp  razor.  If  foreign  matters  are  to  be  removed  from  the  tissue — 
e.g.  the  contents  of  the  gut — do  not  wash  them  with  water,  but  with  a  stream  of  normal  salt 
solution  (p.  xxx).  Place  the  tissues  in  a  relatively  large  quantity  of  the  hardening  fluid,  at 
least  twenty  times  the  bulk  of  the  tissue.  Change  the  fluid  at  the  end  of  the  first,  third, 
seventh,  and  tenth  days.  It  is  better  to  place  the  tissues  in  a  shallow  but  wide  bottle.  See 
that  the  tissues  do  not  stick  to  the  wall  of  the  vessel,  else  the  hardening  fluid  is  prevented 
from  having  access  to  them.  Cotton  wool  moistened  with  dilute  alcohol  answers  admirably 
as  a  bed  for  the  tissues  to  rest  on  and  to  separate  one  piece  from  another.  Place  the 
bottle  in  a  cool  place.  The  hardening  will  be  effected  in  from  one  to  four  weeks.  At  the  end 
of  this  time  remove  the  tissues  ;  wash  them  thoroughly  in  water,  or  let  them  steep  in  a  large 
volume  of  water  for  several  hours  to  get  rid  of  the  hardening  reagent,  as  in  some  cases — e.g. 
chromic  acid — its  presence  interferes  with  the  process  of  staining.  Place  the  tissues  in  equal 
parts  of  methylated  spirit  and  water  for  two  days,  and  then  transfer  them  to  methylated 
spirit  to  complete  the  hardening.  If  the  spirit  becomes  cloudy  it  must  be  changed.  This 
keeps  the  preparations  until  they  are  required  for  sections.  Instead  of  merely  placing  the  tissue 
in  the  hardening  fluid,  in  many  cases  it  is  advisable  to  suspend  it  by  means  of  a  thin  cord  in 
tJie  fluid,  which  permits  the  fluid  to  penetrate  the  tissue  more  rapidly. 

The  process  of  changing  the  various  hardening  fluids  on  certain  days  appears  tedious, 
but  it  is  not  so  in  reality,  if  the  tissues  to  be  hardened  are  placed  on  a  shelf  by  themselves. 
Chromic  acid  preparations  require  to  have  the  fluid  changed  more  frequently  than  when 
Miiller's  fluid  is  employed.  For  convenience,  therefore,  they  may  be  placed  by  themselves. 
On  all  occasions  label  each  bottle,  and  note  the  tissue,  from  what  animal  it  is  taken,  the  name 
and  strength  of  the  hardening  fluid,  and  the  date  on  which  it  was  changed.  Attention  to 
these  apparently  small  details  is  the  necessary  condition  for  successful  histological  work. 


HARDENING   SOLUTIONS. 

The  following  are  the  solutions  in  most  common  use. 

A.      THOSE   WHICH    HARDEN   TISSUES   WITHOUT   COLOURING   THEM. 

1.  Chromic  acid  1  per  cent,  solution. — Dissolve  lo  grms.  chromic  acid  in  i,ooo  c.c.  water. 
So  strong  a  solution  is  not  used,  as  it  renders  the  tissues  too  brittle.  It  is  convenient  to  keep 
a  strong  solution,  which  can  be  diluted  as  required.  A  ;^  or  ^  per  cent,  solution  is  generally 
used. 

2.  Chromic  Acid  and  Spirit  Solution. — This  is  one  of  the  most  useful  solutions.  Make  a 
i  per  cent,  solution  of  chromic  acid,  either  by  diluting  the  above,  or  dissolving  one  gramme  of 
chromic  acid  in  600  c.c. 'of  water.  Take  one  part  of  this,  and  add  two  parts  of  methylated  spirit. 
It  is  better  to  prepare  this  mixture  as  it  is  required.  As  heat  is  developed  by  the  mixing,  the 
tissues  must  not  be  put  into  the  mixture  till  it  becomes  cool.  It  hardens  in  from  seven  to 
ten  days,  according  to  the  size  and  nature  of  the  tissue. 

3.  Potassic  Bichromate  Solution.— Dissolve  20  grms.  in  1,000  c.c.  water.  The  solution  is 
most  easily  made  with  warm  water,  but  it  must  be  allowed  to  cool  before  it  is  used.  If  the 
solution  is  changed  every  four  days  it  hardens  tissues  in  three  to  four  weeks,  but  usually  a 
longer  time  is  required,  as  it  is  not  necessary  to  change  the  fluid  so  often. 


xxxii  INTRODUCTION. 

4.  Ammonium  Bichromate  Solution. — Dissolve  20  grms.  in  1,000  c.c.  water.  This  is  very 
useful  for  hardening  the  brain  and  spinal  cord,  and  nervous  structures  generally. 

5.  Ammonium  Chromate  Solution. — Dissolve  50  grms.  of  ammonium  chromate  in  1,000  c.c. 
water,  i.e.  a  five  per  cent,  solution,  or  it  may  be  made  by  adding  one  ounce  of  the  salt  to 
twenty  fluid  ounces  of  water.  Filter  and  preserve  in  a  stoppered  bottle.  It  hardens  fresh 
tissue,  such  as  the  mesentery,  in  twenty-four  hours.  After  hardening,  the  tissue  must  be  washed 
until  no  more  colour  is  removed  by  the  water.  It  may  then  be  stained  and  mounted  in  gly- 
cerine. For  other  cases,  the  kidney  for  instance,  small  pieces  are  hardened  for  forty-eight 
hours,  and  after  being  thoroughly  washed  are  then  treated  with  spirit,  as  directed  above 
(p.  xxxi).  This  substance  is  of  the  utmost  value  for  revealing  the  rod-like  structure  in  the 
renal  epithelium,  and  for  demonstrating  the  existence  of  the  intra-cellular  and  intra-nuclear 
ple.xus  of  fibrils  in  cells. 

6.  Miiller's  Fluid. — Dissolve  25  grms.  potassic  bichromate  and  10  grms.  sodic  sulphate  in 
1,000  c.c.  water.  This  is  a  most  useful  reagent,  as  it  penetrates  more  readily  than  chromic 
acid,  though  it  takes  longer  time  to  harden  tissues — usually  from  five  to  seven  weeks. 

It  is  frequently  advantageous  to  combine  the  use  of  two  of  the  above  fluids,  thus:  place  the 
tissue  for  a  week  in  Miiller's  fluid,  and  complete  the  hardening  in  No.  2.  After  hardening, 
the  tissues  are  well  washed,  and  preserved  in  spirit. 

7.  Miiller's  Fluid  and  Spirit. —  Mix  3  parts  of  Miiller's  fluid  and  i  part  of  methylated 
spirit.  This  does  very  well  for  nerve-tissues,  muscle,  and  the  retina.  It  ought  to  be  prepared 
fresh  when  required,  and  the  tissue  ought  not  to  be  placed  in  it  till  the  mixture  cools  after  the 
addition  of  the  spirit.  To  prevent  the  separation  of  the  chromium  salts,  which  is  apt  to  occur, 
keep  the  mi.xture  in  a  dark  place. 

8.  Methylated  Spirit  is  sometimes,  though  rarely,  used  alone. 

9.  Absolute  Alcohol  of  S.  G.  0'795. — This  hardens  very  rapidly — in  twenty-four  hours — but 
it  causes  considerable  shrinking,  though  it  is  invaluable  for  gastric  mucous  membrane,  and  for 
secretory  glands  generally,  e.g.  the  salivary  glands  and  pancreas.  Tissues  become  stained  ver>' 
readily  after  hardening  in  pure  alcohol. 


B.      THOSE   WHICH    HARDEN,   AND   AT   THE   .SAME   TIME   COLOUR   THE   TISSUE. 

10.  Picric  Acid. — Make  a  watery  solution  saturated  in  the  cold.  Keep  crystals  in  the 
bottle  to  ensure  saturation.  Tissues  ought  to  be  left  in  this  only  for  a  day  or  two,  else  they 
are  rendered  too  brittle.  Its  stain  is  of  a  bright  yellow  colour,  which  is  easily  removed  by 
prolonged  washing  in  water  (Ranvier).  Tissues  hardened  in  this  fluid  become  easily  stained 
with  picrocarmine. 

1 1.  Kleinenberg's  Picric  Acid  is  a  modification  of  the  above.  To  100  c.c.  of  a  cold  saturated 
watery  solution  of  picric  acid  add  2  c.c.  of  strong  sulphuric  acid,  which  throws  down  a  yellow 
precipitate.  Filter,  and  to  the  filtrate  add  300  c.c.  of  distilled  water.  This  solution  is  most 
valuable  for  fcetal  tissues,  and  especially  for  early  embryos.  It  produces  its  effects  in  from 
three  to  ten  hours. 

1 3.  Osmic  Acid. — A  one  per  cent,  waterj'  solution  is  most  useful,  which  can  be  diluted  as 
required.  A  ^  per  cent,  solution,  prepared  with  distilled  water,  is  usually  employed.  It 
decomposes  very  rapidly  when  in  contact  with  organic  matter  and  exposed  to  light ;  and  as 
it  is  very  volatile,  it  must  be  kept  in  a  glass-stoppered  bottle,  covered  with  paper,  to  protect  it 
from  the  light.  It  hardens  in  from  six  to  twenty-four  hours,  and  acts  specially  on  fatty 
matters,  which  it  blackens,  and  is  very  useful  for  tracing  the  course  of  medullated  nerve-fibres, 


INTRODUCTION.  xxxiii 

as  it  blackens  the  myeline.  Small  pieces  of  tissue  exposed  to  the  vapour  of  a  one  per  cent, 
solution  for  a  few  minutes,  are  easily  stained  with  picrocarmine,  though  the  prolonged  action 
of  a  strong  solution  renders  subsequent  staining  difficult.  It  is  one  of  the  most  useful  of 
all  hardening  reagents  (M.  Schultze).  It  is  not  necessary  to  mount  preparations  treated  with 
osmic  acid  in  a  saturated  solution  of  acetate  of  potash. 


METHODS   OF   SOFTENING   TISSUES. 

Certain  tissues,  as  bone  and  tooth,  which  contain  calcareous  matter,  require  to  have  the 
calcareous  matter  removed  before  they  can  be  conveniently  cut  for  the  microscope.  This  is 
done  by  placing  them  in  a  decalcifying  solution.  Some  solutions  remove  only  the  calcareous 
matter  —e.g.  dilute  hydrochloric  acid — without  at  the  same  time  hardening  the  tissue;  others,  as 
a  mixture  of  chromic  acid  and  nitric  acid,  remove  the  lime  salts,  and  at  the  same  time  harden 
the  tissue,  and  generally  the  latter  are  to  be  preferred. 


DECALCIFYING   SOLUTIONS. 

1.  Hydrochloric  Acid. — Mix  one  part  of  strong  acid  with  ten  parts  of  water.  This  does 
very  well  for  the  removal  of  lime  salts  from  an  injected  bone. 

2.  Chromic  and  Nilsric  Acid  Fluid. — Make  a  \  per  cent,  solution  of  chromic  acid,  and  to 
every  lOO  c.c.  add  i  c.c.  of  strong  nitric  acid.  The  chromic  acid  solution  is  easily  made  by 
adding  an  equal  volume  of  water  to  a  one  per  cent,  solution  (p.  xxxi).  This  fluid  takes  two  or 
three  weeks  to  remove  the  lime-salts  from  a  small  piece  of  bone,  but  if  the  fluid  is  frequently 
changed  the  result  is  accomplished  more  rapidly.  The  tissue  is  then  thoroughly  washed  to 
get  rid  of  the  acids,  and  then  hardened,  first  in  weak  and  then  in  strong  methylated  spirit. 
The  advantage  of  this  fluid  is  that  the  chromic  acid  hardens  the  parts,  whilst  the  nitric  acid 
removes  the  lime-salts.  Bones,  after  being  exposed  to  its  action,  assume  a  green  colour,  owing 
to  the  formation  of  a  sesquioxide  of  chromium. 

3.  Picric  Acid  Solution. — A  saturated  watery  solution  takes  several  weeks  to  act  on  a 
moderately  large  piece  of  bone.     It  is  of  great  value  for  fcetal  bones. 

4.  A  10  per  cent.  Solution  of  Common  Salt  and  Hydrochloric  Acid. — This  is  most  valuable 
for  showing  the  matrix  of  bone,  which  consists  of  ordinary  fibrous  tissue,  and  swells  up  in  the 
ordinary  acid  media.  A  10  per  cent,  solution  of  salt  prevents  this  (v.  Ebner  and  De  B.  Birch). 
The  bone  is  placed  in  a  10  per  cent,  solution  of  common  salt,  to  which  i  to  3  per  cent,  of 
hydrochloric  acid  is  added.  Add  from  day  to  day  as  much  acid  as  will  decalcify  the  bone. 
When  the  bone  becomes  flexible  It  is  placed  for  several  hours  in  water,  to  remove  all  the  acid. 
Leave  it  for  several  days  in  10  per  cent,  salt  solution,  which  must  be  changed  repeatedly. 
When  the  reaction  of  the  bone  becomes  neutral  the  bone  is  white  and  opaque.  Sections  are 
made,  and  mounted  in  a  10  per  cent,  solution  of  salt.  They  show  the  fibrillar  structure  of  the 
matri.x  (p.  32)  (v.  Ebner). 


xxxiv  INTRODUCTION. 


METHODS    FOR  DISSOCIATING   TISSUES. 

Various  solutions  dissolve  or  soften  certain  parts  of  a  tissue  whilst  other  parts  are  left  un- 
affected. The  result  is  that  the  component  parts  may  be  readily  separated  by  teasing.  The 
piece  of  tissue  ought  not  to  be  larger  than  a  pea.  The  result  is  usuall)' effected  in  from  t\\'cnt}-- 
four  to  thirty-six  hours,  though  much  less  time  may  suffice. 


DISSOCIATING   SOLUTIONS. 

1.  Iodised  Serum. — Add  iodine  to  blood-serum  or  amniotic  fluid,  till  the  fluid  is  of  a  dis- 
tinctly yellow  colour.  This  fluid  dissolves  the  cement-substance  between  cells  in  from  one  to 
two  days. 

2.  Dilute  Chromic  Acid  (-oi  per  cent.). — Dissolve  i  grm.  chromic  acid  in  10,000  c.c.  water, 
or  dilute  a  one  per  cent,  solution.  This  does  excellently  for  isolating  the  fibriUa;  of  muscle,  and 
for  the  nerve-cells  of  the  spinal  cord.  Two  to  three  daj-s'  maceration  serves  to  bring  about  the 
result. 

3.  Dilute  Alcohol  ('  Alcool  au  tiers  '). — Mix  2  parts  of  water  with  i  of  rectified  spirit.  This 
is  one  of  the  most  useful  dissociating  fluids,  and  requires  one  to  tv/o  days  for  its  action 
(Ranvier). 

4.  Saturated  Aqueous  Solution  of  Baric  Hydrate  requires  about  twenty-four  hours  to  act  on 
the  fibrillae  of  tendon. 

5.  Caustic  Potash. — Dissolve  40  grms.  of  caustic  potash  in  100  c.c.  water.  This  isolates 
muscle-cells  in  from  twenty  to  thirty  minutes. 

6.  Ten  per  cent.  Solution  of  Common  Salt  is  useful  for  dissolving  the  cement  of  white  fibrous 
tissue.  It  takes  several  days  to  act.  It  is  very  useful  also  for  showing  the  fibrillse  of  the 
matrix  of  bone. 

7.  Nitric  Acid  and  Glycerine. — Mix  one  part  of  strong  nitric  acid  containing  nitrous  acid 
with  3  parts  of  water  and  i  part  of  glycerine.  The  object  is  placed  in  this  mixture  for  two 
or  three  days  and  then  in  water.  It  is  specially  useful  for  isolating  nerve-structures  and  lens- 
fibres  (Freud). 

DIGESTION    AS   A    HISTOLOGICAL    METHOD. 

This  method  was  introduced  into  histology  by  the  author  several  years  ago.  It  has  re- 
cently been  employed  by  Kuhne  for  investigating  the  structure  of  nerves,  and  by  De  Burgh 
Birch  in  his  investigations  on  the  composition  of  the  matrix  of  bone  (p.  32).  Either  an  arti- 
ficial gastric  or  pancreatic  juice  may  be  employed. 

Artificial  Gastric  Digestion. — This  method  is  fully  described  at  p.  92  (W.  Stirling). 

Artificial  pancreatic,  i.e.  trypsin,  Digestion. — Either  an  aqueous  or  glycerine  extract  of  the 
pancreas  may  be  used.  The  latter  is  the  more  convenient.  It  is  made  thus  by  v.  Wittich's 
method.  The  pancreas  of  a  dog  is  chopped  up  and  is  dehydrated  with  absolute  alcohol  for 
twenty-four  hours.  The  alcohol  is  removed  and  sufficient  pure  glycerine  is  added  to  cover 
the  gland,  and  it  is  allowed  to  stand  for  three  weeks.  Press  the  glycerine  through  muslin  to 
remove  the  gland  tissue.    The  glycerine  is  a  solvent  for  the  trypsin  of  the  pancreas,  just  as  it  is 


INTRODUCTION. 


XXXV 


for  other  soluble  ferments,  i  c.c.  of  the  glycerine  filtrate  is  added  to  19  c.c.  of  1  per  cent, 
solution  of  sodic  carbonate.  The  fluid  becomes  turbid,  but  after  filtration  a  pale  yellow  fluid 
is  obtained.  The  tissue  to  be  digested  is  placed  in  this  fluid,  and  the  whole  is  kept  at  a  tem- 
perature of  40°  C.  in  a  water  bath  as  directed  at  p.  92.  Sections  of  softened  bone  digested  by 
this  method  are  preserved  in  a  10  per  cent,  solution  of  common  salt  (Birch). 


METHODS    OF   TEASING   A   TISSUE. 

Take  only  a  small'  piece  of  tissue,  and  place  it  on  a  slide  in  a  small  drop  of  the  fluid  in 
which  it  is  to  be  mounted,  e.g.  glycerine.  Fix  one  end  of  the  tissue  with  a  strong  needle  in  a 
handle,  and  tear  in  the  direction  of  the  fibres  with  the  other  needle.  The  low  power  must  be 
used  from  time  to  time,  to  ascertain  when  the  component  parts  are  sufficiently  isolated.  It  is 
very  desirable  to  have  a  proper  background,  so  that  the  object  to  be  teased  may  be  distinctly 
seen  ;  for  a  coloured  object  a  small  piece  of  white  bibulous  paper  answers  admirably,  and  a  trans- 
parent object  is  generally  best  seen  on  a  dark  surface.  A  good  plan  is  to  have  a  narrow  white 
line  painted  along  the  edge  of  the  work-table.  The  same  result  is  obtained  by  having  a  small 
slab,  one-half  of  which  is  glazed  white  and  the  other  black. 


Into 


Fig. 


15.    Ranvier's  Photophokf.. 
One-si.\th  Natural  Size. 


r.a.nvier's  photofhore. 

The  following  little  arrangement  devised  by  Ranvier  is  of  great  value  for  this  pur- 
pose. It  is  called  a  photophore  (fig.  15).  It  consists  of  a  small  mahogany  box  about  four 
inches  square  ;  one  side  is  open  and  so  is  the  top.  The  lid  is  formed  of  a  glass  plate, 
the  bo.x  a  mirror  is  fixed  at  an  angle  of  30°-3S°.  This  arrange- 
ment reflects  the  light  through  the  preparation.  If  it  be  desired 
to  work  with  a  black  background,  all  that  is  required  is  to  place 
a  black  card  over  the  mirror.  This  simple  arrangement  is  of 
the  utmost  value,  and  when  in  use  has  simply  to  be  directed  with 
its  mirror  towards  a  window.  It  is  well  to  have  a  bent  piece 
of  wire,  with  a  ring  formed  on  it  and  attached  to  the  back  of  the 

box,  for  receiving  a  small  magnifying  lens.  Each  student  should  be  provided  with  such  an 
arrangement.  He  can  easily  make  one  for  himself  by  taking  an  ordinary  small  cigar-box 
without  a  lid,  and  placing  in  it  a  mirror  at  an  angle  of  SO'-SS".  On  the  upper  side  make 
a  square  hole,  and  into  this  fit  a  glass  plate. 


SIMPLE   OR   DISSECTING   MICROSCOPE. 

In  teasing  or  dissociating  a  tissue  it  is  often  of  great  value  to 
use  an  ordinary  dissecting  microscope.  The  form  devised  by  Bri.ickc 
and  made  by  Hartnack  is  useful,  but  it  is  expensive.  For  practical 
purposes  I  find  that  the  form,  fig.  16,  made  by  Swift,  is  a  most  useful 
and  handy  instrument. 


Fig.    16.     Swift's  Si.mi'i.e 

Dissecting  Microscope. 

One-sixth  Natural  Size. 


d2 


xxxvi  INTRODUCTION. 


METHODS  OF  CUTTING  SECTIONS. 

It  is  of  the  utmost   importance  that   the   student  should   thoroughly  master  the  details  of 
cutting  sections  by  hand,  and  also  with  the  aid  of  certain  instruments  called  microtomes. 

I.  SECTIONS   OF   UNHARDENED   TISSUES. 

If  it  be  desired  to  examine  only  a  small  piece,  snip  off  a  thin  fragment  with  a  pair  of 
scissors  curved  on  the  flat,  or  cut  off  a  slice  with  a  Valentine's  knife. 

II.  SECTIONS    OF    HARDENED    TISSUES. 

I.  Cutting  sections  by  hand  with  a  razor. — Tissues  hardened  by  any  of  the  above-mentioned 
methods  are  difficult  to  cut  unless  they  are  clamped  or  imbedded  in  some  easily  cut  material, 
which  must  have  nearly  the  same  consistence  as  the  tissue  itself  As  to  the  kind  of  razor  or 
knife  to  employ,  the  '  army  razor  '  answers  admirably,  though  some  prefer  specially  constructed 
knives,  which,  I  believe,  possess  no  advantages,  whilst  others  prefer  the  razor  to  be  ground 
flat  on  the  side  directed  towards  the  tissue  to  be  cut. 

If  the  piece  of  tissue  be  sufficiently  large,  seize  it  between  the  thumb  and  fore-finger  of  the 
left  hand.  Hold  the  fore-finger  horizontally,  so  that  its  outer  surface  forms  a  rest  on  which 
the  blade  of  the  razor  may  glide.  Keep  the  handle  of  the  razor  in  a  line  with  the  blade,  and 
hold  it  firmly  in  the  right  hand.  Draiv  it  (from  heel  to  tip)  through  the  tissue  towards 
yourself  Keep  the  razor  ivcll  ivetted  ivitli.  spirit,  and  float  the  section  off  into  spirit  or  water, 
with  a  camel-hair  pencil.  It  requires  very  considerable  practice  before  one  becomes  expert  at 
making  sections  of  sufficient  thinness  to  be  of  use.  The  razor  must  be  kept  perfectly  sharp, 
and  after  making  a  few  sections  should  always  be  '  stropped.' 

If  the  tissue  be  too  small  or  too  delicate  to  hold  in  the  hand,  it  must  either  be  clamped  or 
imbedded  in  some  substance. 

{a)  A  piece  of  amyloid  or  waxy  liver,  or  the  liver  of  a  pig  hardened  in  alcohol,  serves  the 
purpose  admirably.  Make  a  slit  in  the  piece  of  liver,  and  clamp  the  tissue  in  it,  and  cut  both 
liver  and  tissue  as  directed  above. 

(J?)  Carrot  or  elder-pith  may  be  used  instead.  Make  a  slit  in  the  carrot  or  pith,  and  clamp 
in  it  the  tissue  to  be  cut. 

IMBEDDING   MIXTURES. 

{a)  Melt  together  with  the  aid  of  a  gentle  heat  four  parts  of  ordinary  solid  paraffine  with 
one  of  pure  lard.  A  quantity  of  this  mixture  should  be  kept  ready  for  use.  When  required, 
the  mixture  is  to  be  dissolved  in  a  vessel  heated  in  a  water-bath  at  the  lowest  temperature 
possible. 


INTRODUCTION. 


XXXVll 


Method  of  Imbedding. — Press  the  tissue  to  be  imbedded  between  folds  of  blotting-paper, 
to  remove  all  the  alcohol,  and  see  that  its  surface  is  thoroughly  Axy.  It  is  a  good  plan  to  dip 
it  for  a  moment  into  a  weak  solution  of  gum  arable,  and  allow  the  gum  to  dry  as  a  film  on 
its  surface.  This  prevents  the  paraffin  from  penetrating  the  tissue,  and  also  causes  it  to 
separate  more  readily  from  the  section.  Fold  a  piece  of  paper  in  the  form  of  an  oblong  box, 
and  pour  into  it  a  little  of  the  previously  melted  paraffine.  Place  a  needle  into  the  tissue,  fix 
it  conveniently  in  the  paraffine  mixture  in  the  paper  box,  add  more  paraffine  until  the  tissue  is 
well  covered.  Use  the  needle  to  keep  the  tissue  in  its  place  until  the  mixture  sets.  The 
paraffine  block  is  then  removed  from  the  paper,  and  sections  cut  through  it  and  the  imbedded 
tissue.     The  sections  are  floated  off  into  spirit  or  water. 

{J})  Other  imbedding  mixtures  are  sometimes  used,  as  two  parts  of  paraffine  mixed  with 
one  of  vaseline,  which  gives  a  transparent  mass,  in  which  the  position  of  the  tissue  can  be 
detected. 

MICROTOMES. 

When  a  large  number  of  sections  is  required,  or  when  a  complete  section  is  wanted  through 
an  organ,  there  can  be  no  doubt  that  a  microtome  of  some  kind  is  required.  One  of  the  earliest 
microtomes  introduced  (fig.  17)  is  that  of  Mr.  A.  B.  Stirling,  of  the  Anatomical  Museum, 
Edinburgh  University.  It  consists  of  a  strong  brass  tube,  with  a  broad,  smooth,  metal  plate 
fi.xed  at  right  angles  to  it  at  one  end.  In  this  cylinder  a  plug  is  moved  by  a  very  fine- 
threaded  screw,  which  works  in  the  lower  end  of  the  tube.  The  instrument  is  so  made  that  it 
can  be  clamped  to  a  table.  The  tissue  to  be  cut  is  imbedded  in  the  well  of  the  microtome  in 
paraffine  and  lard,  in  the  same  way  as  a  tissue  is  imbedded  for  cutting  hand  sections.  Melt 
the  imbedding  mi.xture,  and  fill  the  well  of  the  microtome,  and  whilst  still  fluid  put  in  the 
tissue  to  be  cut — using  the  same  precautions  as  to  drj-ing — and  hold  it  in  position  till  the 
mixture  sets.  By  turning  the  screw  the  mass  and  included  tissue  are  elevated  ;  and  a  sharp  knife 
carried  along  on  the  flat  brass  plate  shaves  off  a  section,  which  can  be  made  as  thin  as  desired 
by  having  a  screw  of  sufficient  fineness.  The  knife  must  be  kept  flooded  with  spirit,  and  the 
sections  are  floated  off  by  means  of  a  camel-hair  brush  into  spirit  or  water. 


Fig.  17.    Mr.  a.  B.  Stirling's  Microtome. 


Fig.  iS.    Ranvier's  Hand  Microtome. 
Half  Natural  Size. 


Ranvier's  Microtome  is  simply  a  smaller  instrument,  working  on  the  same  principle,  and 
designed  to  be  held  in  the  hand  (fig.  18).  The  whole  apparatus  can  be  inverted  in  spirit,  if  it 
be  desired  to  make  sections  of  the  same  tissue  from  time  to  time. 


XXXVlll 


INTRODUCriON. 


Sections  to  be  cut  in  Ranvier's  microtome  are  imbedded  just  as  in  Mr.  Stirling's  form, 
with  a  paraffinc  and  lard  or  other  mixture.  Elder-pith,  however,  may  be  used.  The  tissue  is 
packed  in  with  dry  elder-pith,  and  then  the  whole  apparatus  is  placed  in  spirit,  which  causes 
the  pith  to  swell,  and  so  to  fix  the  tissue  sufficiently  firmly  for  a  section  to  be  made  of  it. 


FREEZING    MICROTOMES. 

Dr.  Rutherford's  Freezing  Microtome. — This  microtome  (fig.  19)  is  essentially  a  modification 
of  Mr.  Stirling's  microtome.  The  hollow  brass  cylinder  is  surrounded  with  a  large  trough  (g), 
which  is  filled  with  a  freezing  mixture  composed  of  equal  parts  of  salt  and  ice.  The  trough  is 
provided  with  a  tube  (h)  to  permit  of  the  outflow  of  the  water  resulting  from  the  melting  of 
the  ice.  It  is  a  great  improvement  to  have  the  cutting- 
table  covered  with  a  thick  plate  of  glass,  as  this  is  not  worn 
away  so  readily  as  the  brass.  This  microtome  can  be  used 
as  an  ordinary  microtome,  as  well  as  for  freezing. 

METHOD   OF   USING   A  FREEZING   MICROTOME. 

Preparation  of  the  Microtome. —  Clamp  the  instrument 
to  a  table.  Turn  the  screw  (d)  until  the  plug  has  descended 
a  good  distance  into  the  well,  and  to  prevent  the  screw 
freezing  pour  a  little  alcohol  into  the  well  and  let  it  run 
through  the  tube  ;  dry  the  well  thoroughly,  and  smear  a 
little  lard  round  the  top  of  the  plug,  to  prevent  the  escape 
of  any  of  the  imbedding  mixture  between  the  plug  and 
the  wall  of  the  cylinder,  which  would  interfere  with  the 
action  of  the  screw. 

Preparation  of  the  Tissue  to  be  Frozen. — The  tissue  has 
first  been  properly  hardened  by  one  of  the  foregoing 
methods.      If   it   has  been    kept    in    alcohol,    it    must    be 

placed  in  a  large  quantity  of  water — to  be  changed  several  times — during  twenty-four  hours, 
to  remove  all  the  alcohol.  It  is  then  placed  for  twenty-four  hours  in  gum-mucilage  (B.  P.).  The 
well  is  then  filled  with  gum  (B.  P.),  and  equal  quantities  of  powdered  ice  and  salt  are  packed 
into  the  trough  with  a  piece  of  stick  until  the  edge  of  the  gum  is  observed  to  freeze,  when  the 
piece  of  tissue  is  placed  in  the  gum  in  the  side  of  the  well  nearest  the  side  from  which  the 
person  makes  the  sections,  by  means  of  a  needle.  The  freezing  process  is  continued  until  the 
whole  mass  of  gum  becomes  solid,  the  gum  becoming  of  such  a  consistence  that  it  cuts  like  a 
piece  of  cheese  (U.  Pritchard). 

The  Section  Knife. — The  ordinary  razor  does  not  answer  well  for  cutting  sections  with  the 
microtome.  Its  blade  is  too  thin  and  too  short.  It  is  necessary  to  have  a  stout  blade,  seven 
or  eight  inches  long,  with  a  tJiick  straight  back — at  least  a  quarter  of  an  inch  thick — fixed  in 
a  straight  handle-.  The  blade  must  be  so  strong  that  it  cannot  be  bent  whilst  pushing  it 
along  the  glass  plate.  The  surface  next  the  glass  plate  ought  to  be  slightly  concave.  The 
surface  of  the  knife  ought  to  be  moistened  with  gum  solution,  which  prevents  the  sections 
from  curling  up.  In  cutting,  keep  the  back  of  the  knife  directed  towards  the  operator,  and 
push  the  blade  obliquely  from  tip  to  heel,  guiding  it  with  the  thumb  and  forefinger  of  the  left 
hand  through  the  tissue.  Each  section  as  it  is  made  must  be  floated  off  into  a  bowl  of  water 
with  the  aid  of  a  large  camel-hair  brush.     (See  p.  xl  for  how  to  preserve  them.) 


Fig.  19.  Dr.  Ruthekkord's  Freezing 
Microtome. 

B,  Plate  of  brass  with  aperture  A  leading 
into  the  well  of  the  Microtome;  c,  Lower 
end  of  cylinder  in  which  the  screw  D 
works,  moving  a  brass  plug  up  or  down  ; 
G,  Trough  to  hold  freezing  mixture  ;  n. 
Exit  tube  for  water;  F,  Clamp  to  fix  the 
instrument  to  a  table. 


INTRODUCTION. 


XXXIX 


MODIFICATIONS   OF   THE   ABOVE   METHOD. 

In  the  preparation  of  tlie  Tissues  (a). — For  some  tissues — e.^.  brain — it  is  necessary  to  leave 
them  forty-eight  hours  in  the  gum  before  cutting. 

(/;)  The  following  method  is  an  excellent  one,  and  is  of  the  greatest  value  for  very  delicate 
tissues  as  the  retina,  ear,  brain.  After  soaking  the  tissues  in  water,  place  them  for  twenty- 
four  hours  in  syrup  made  by  dissolving  four  ounces  of  crystallised  sugar  in  two  ounces  of 
water.  They  are  afterwards  placed  in  gum  for  twenty-four  hours,  and  are  then  read}'  for 
cutting. 

With  such  a  microtome  as  described  above  there  is  no  difficulty  whatsoever  in  making  the 
most  perfect  sections.  The  well  can  be  made  of  any  size,  and,  indeed,  D.  J.  Hamilton,  of 
Edinburgh,  has  had  a  large  freezing  microtome  constructed  which  enables  him  to  make 
microscopic  sections  of  an  entire  brain. 


\VILLIAM.S'   FREEZING   MICROTOME. 

Some  histologists  prefer  this  form  of  microtome  (fig.  20).  It  is  made  by  Swift,  of  London, 
and  consists  of  a  wooden  tub  for  the  freezing  mixture.  In  this  is  an  upright,  brass  bar,  into 
whose  upper  end  the  circular  brass  plates,  on  which  the  tissue  is  frozen,  are  screwed.  A  glass 
top  with  a  hole  in  its  centre,  through  which  the  circular  plate  projects,  forms  a  lid  for  the  box. 
The  knife  is  fixed  in  a  triangular  frame,  provided  with  screws  by  which  it  can  be  raised  or 
lowered. 

METHOD   OF   USING   WILLIAM.?'   MICROTOME. 

Remove  the  lid  of  the  box  and  fill  the  chamber  with  equal  parts  of  pulverised  ice  and  salt, 
care  being  taken  not  to  allow  the  mixture  to  touch  the  under  side  of  the  cover,  which,  when 


Fig.  20.   Williams'  (Quecket  Club)  Freezing  Microtom^. 

replaced,  must  be  firmly  secured  by  the  clamp  screw  for  that  purpose.  The  substance  to 
be  cut  is  placed  on  the  surface  of  the  central  circular  brass  piece,  and  surrounded  with  a 
solution  of  gum,  which  readily  congeals,  and  will  thus  hold  the  specimen   firmly  in   position, 


xl  INTRODUCTION. 

which  will  solidify  shortly  after  the  gum  has  frozen.  It  is  advisable  to  cover  the  apparatus 
with  baize,  to  facilitate  the  freezing  process.  The  edge  of  the  razor  must  be  elevated  to  the 
required  height  for  cutting  the  section  by  means  of  the  three  screws  supporting  the  frame. 
After  the  first  cut,  each  end  of  the  razor  must  be  again  presented  to  the  surface  of  the 
specimen,  when  either  end  of  the  blade  can  be  adjusted  by  one  of  the  back  screws  until  its 
entire  length  is  level  ;  then  by  turning  the  large  screw  in  the  front  part  of  the  frame  it  can  be 
lowered  for  each  successive  section  required. 

Preparation  of  the  Material  for  Freezing. — Follow  exactly  the  same  methods  as  are  in- 
dicated at  p.  xxxviii.  A  piece  of  tissue,  a  quarter  of  an  inch  in  thickness,  is  quite  sufficient 
to  freeze  at  one  time.  The  tissue  to  be  frozen  is,  after  the  above  preparation,  placed  on  one 
of  the  circular  plates,  and  is  fixed  to  it  with  gum  solution.  The  razor  ought  to  be  moistened 
with  the  same  solution,  which  prevents  the  sections  from  curling  up. 

TREATMENT   OF   THE   SECTIONS. 

From  the  bowl  of  water,  into  which  they  are  put  at  first,  they  are  transferred  to  a  conical 
glass  filled  with  water,  in  which  they  gradually  subside.  All  the  gum  must  be  got  rid  of,  which 
is  accomplished  by  changing  the  water  several  times.  When  all  the  gum  is  dissolved,  transfer 
the  sections  to  one  or  other  of  the  following  fluids  till  they  are  required.  Keep  them  in  glass- 
stoppered  bottles. 

PRESERVATIVE   FLUIDS. 

1.  Ordinary  methylated  spirit. 

2.  Glycerine  i  oz.,  water  i  oz.,  carbolic  acid  4  minims. 

DR.   BEVAN    lewis's   ETHER   FREEZING    MICROTOME. 

It  is  not  always  possible  to  obtain  ice  for  freezing,  and  hence  it  is  necessary  to  have 
another  medium  for  freezing.  This  is  supplied  in  the  most  perfect  form  by  the  above  in- 
strument (fig.  21),  devised  by  Dr.  Bevan  Lewis,  of  the  West  Riding  Asylum.  This  microtome 
is  specially  valuable  for  rapid  freezing. 

The  lower  half  or  body  of  the  instrument  {a)  is  a  slightly  modified  Stirling  microtome; 
the  upper  part  {p)  consists  of  a  freezing-chamber  and  a  section-plate. 

The  Body.—  'Y\\&  framework  of  the  body  {a)  is  a  solid  brass  casting,  which  can  be  secured 
to  a  table  firmly  by  the  coarse  screw  (w).  Through  its  central  aperture  works  the  brass  plug 
{li),  driven  by  a  spring  and  fine  micrometer  screw  ik),  the  pitch  being  fifty  threads  to  the  inch, 
the  screw  having  a  diameter  of  ^\ths  of  an  inch  with  a  milled  head  i^  inches  across. 

Section-Plate. — The  section-plate  {c)  is  a  circular  plate  of  zinc  f  ths  of  an  inch  thick,  raised 
upon  a  strongly  vertical  arm  of  brass  or  zinc — preferably  of  the  latter  metal.  In  the  section- 
plate  there  is  a  circular  central  aperture  through  which  the  freezing-chamber  works.  Lately 
the  efficiency  of  the  instrument  has  been  much  improved  by  capping  the  zinc  plate  [c]  with  a 
square  of  plate-glass  |-inch  thick,  extra  polished  and  drilled  with  a  central  aperture  («)  corre- 
sponding in  size  to  that  in  the  zinc  plate,  and  which  in  my  own  instrument  measures  2^  inches 
in  diameter.  For  cutting  sections  of  frozen  tissue,  the  glass-section  plate  is  a  great  advantage, 
and  when  once  used  will  never  be  dispensed  with.  It  renders  the  movements  of  the  knife 
absolutely  free  and  easy,  whilst,  on  the  other  hand,  the  blade  never  gets  injured  from  scratches 
which  sooner  or  later  invariably  appear  upon  a  metal  section-plate,  and  which  turn  or  indent 
the  edge. 


INTRODUCTION.  xli 

Freezing-Chamber. — This  consists  of  a  zinc  cylinder  ip)  \\  inches  deep  by  2\  wide,  closed 
above  and  below.  An  aperture  (/),  |  of  an  inch  across,  is  made  on  either  side  of  the  cylinder 
to  allow  the  admission  of  the  nozzle  of  the  spray-producer,  as  also  for  free  evaporation  of 
ether,  a  large  proportion  of  which,  however,  condenses  on  the  sloping  false  bottom  i^g)  and  is 
conveyed  off  by  the  tube  (/)  into  a  bottle  attached  to  it. 


Fig.  21.    The  Ether  Freezing  Microtome  of  Dr.  Bevan  Lewis.     This  instrument  is  made  by 
Mr.  Gardner,  South  Bridge,  Edinburgh. 


The  instrument  is  simple,  efficient,  and  compact,  and  enables  one  with  a  minimum 
expenditure  of  ether  to  obtain  the  most  delicate  sections  of  any  tissue  not  more  resistant  than 
cartilage. 

A  little  experience  in  the  use  of  the  instrument  enables  the  operator  to  freeze  so  rapidly 
that  he  will  scarcely  feel  that  it  repays  him  to  collect  the  condensed  ether. 


xlii  INTRODUCTION. 


METHODS   OF  STAINING    TISSUES. 

Tissues  may  be  stained  with  certain  reagents,  e.g.  eosin,  which  give  a  uniform  colour  to  the 
section,  whilst  other  substances  derive  their  value  from  staining  different  parts  of  a  section 
unequally  ;  nuclei,  for  example,  are  usually  stained  of  a  deeper  red  than  the  surrounding 
protoplasm.     For  the  method  of  transferring  large  sections  to  a  slide,  see  p.  xxvi. 

GENERAL    DIRECTIONS. 

It  is  always  advisable  to  use  rather  a  weak  solution  of  the  staining  agent,  as  this  stains 
slowly  and  gives  the  dye  time  to  completely  permeate  the  section.  When  practicable  the 
sections  should  remain  in  the  fluid  for  several  hours.  The  proper  depth  of  tint  can  easily  be 
ascertained  by  lifting  out  a  section  and  washing  it  in  water,  and  examining  it  on  a  slide  with 
a  low  power.  A  little  practice  soon  enables  one  to  tell  when  the  staining  is  deep  enough. 
For  class  purposes,  however,  it  is  often  necessary  to  use  strong  solutions  of  the  dyes,  and 
the  results  are  very  satisfactory. 

For  class  purposes  staining  with  a  strong  dye  is  readily  accomplished.  Float  a  section  on 
to  a  slide  (p.  xxvi),  remove  all  the  water  from  the  slide  and  around  the  section.  Place  on  the 
section  a  large  drop  of  the  staining  fluid,  and  leave  it  'until  the  section  is  properly  stained. 
Wash  off  the  surplus  pigment  in  a  bowl  of  water,  float  the  section  on  to  the  slide  again,  and 
mount  it  in  glycerine  or  other  mounting  fluid. 

STAINING   SOLUTIONS. 

LOGWOOD   SOLUTIONS. 

{a)  A.  Dissolve  '3  grms.  of  haematoxylin  in  lo  c.c.  of  absolute  alcohol.  B.  Dissolve  '3 
grms.  of  alum  in  lOO  c.c.  water.  A  few  drops  of  A  added  to  a  few  c.c.  of  B  gives  a  solution 
of  a  beautiful  violet  colour,  which  rapidly  stains  tissues. 

{b)  A  strong  infusion  of  logwood  chips  is  made  ;  to  this  reddish-coloured  fluid,  after  filtra- 
tion, alum  is  added,  until  a  bluish-violet  solution  results.  The  violet  colour  deepens  by  ex- 
posure to  the  air  for  several  days. 

(c)  Take  60  grms.  of  extract  of  hEematoxylin  and  180  grms.  of  alum.  Rub  them  toge- 
ther in  a  mortar  and  slowly  add  280  c.c.  distilled  water.  Filter,  and  to  the  filtrate  add  20  c.c. 
of  absolute  alcohol.  Keep  all  of  the  above  in  glass-stoppered  bottles.  Logzvood  solutions  ought 
always  to  be  filtered  before  they  are  used. 

I  have  found  solutions  prepared  by  any  of  the  above  methods  to  give  excellent  results. 
They  act  very  rapidly  on  nuclei,  giving  them  a  bright  violet  colour,  softer  and  pleasanter  to 
the  eye  than  the  bright  red  stain  of  carmine.     Many,  however,  prefer  a  solution  which  contains 


INTRODUCTION.  xliii 

alcohol  ;  such  a  solution  is  the  following,  which  has  the  advantage  of  not  requiring  to  be  filtered 
so  frequently. 

id)  Kleinenbergs  Logzvood  Solution. — Make  a  saturated  alcoholic  (70  per  cent.)  solution  of 
crystallised  calcic  chloride,  and  a  similar  solution  of  alum.  Add  8  parts  of  the  alum  solution 
to  I  of  the  calcic  chloride.  To  this  mixture  add,  drop  by  drop,  a  saturated  solution  of 
haemato.xylin  in  absolute  alcohol,  until  the  whole  fluid  assumes  a  deep  purple  colour.  Use 
only  a  small  quantity  of  alcohol,  as  the  haamatoxylln  is  very  soluble.  The  colour  deepens  and 
improves  by  keeping.  When  it  is  desired  to  stain  a  section,  this  solution  must  be  diluted  by 
adding  a  few  drops  of  it  to  a  watch-glass  full  of  water. 

{e)  Logivood  solution  foi-  rapid  staining. — This  solution,  devised  by  E.  A.  Cook,  yields  good 
results,  but  it  has  no  advantages  over  any  of  the  above.  The  ingredients  are  extract  of  log- 
wood and  alum,  of  each,  6  parts,  cupric  sulphate  i  part,  and  water  40  parts.  Grind  in  a  mortar 
the  alum,  logwood,  and  cupric  sulphate,  all  of  which  must  be  iron  free,  and  when  powdered  add 
sufficient  water  to  form  a  thin  paste.  Leave  this  for  one  or  two  days,  then  add  the  remainder 
of  the  water  and  filter.  To  the  filtrate  add  a  crystal  of  thymol,  to  preserve  it  from  mould. 
For  tissues  hardened  in  chromic  acid  add  8  drops  of  this  solution  to  120  drops  of  water,  and 
add  one  drop  of  a  tenth  per  cent,  solution  of  potassic  bichromate  just  prior  to  use. 


CARMINE   SOLUTIONS. 

Strong  Carmine  Solution  for  rapid  staining. — Rub  up  two  grms.  of  the  best  carmine  in  a 
mortar  with  a  few  drops  of  water,  add  4  c.c.  of  liquor  ammoniae  to  dissolve  the  carmine,  then 
add  60  c.c.  of  water.  Filter  the  solution  and  keep  it  in  a  stoppered  bottle.  The  solution  may 
be  diluted,  if  it  be  desired  to  stain  a  tissue  slowly. 

Carmine  is  not  employed  so  frequently  as  it  used  to  be.  A  strong  solution,  such  as  the 
above,  is  apt  to  give  a  somewhat  uniform  tinge  to  the  section.  Better  results  are  obtained  by 
using  either  the  above  fluid  diluted,  or  the  following : — 

Weak  Carmine  Solution. — Rub  up  i  grm.  of  best  carmine  in  a  mortar  with  a  few  c.c.  of 
distilled  water  ;  add  3  c.c.  of  liquor  ammonia,  and,  when  the  carmine  is  dissolved,  add  1 50  c.c. 
of  distilled  water,  and  filter  into  a  stoppered  bottle.  Add  30  c.c.  of  glycerine  and  100  c.c.  of 
rectified  spirit.     This  is  a  modification  of  Beale's  carmine  fluid. 

All  carmine-stained  preparations,  after  being  thoroughly  washed  in  water,  are  improved 
by  placing  them  for  a  few  minutes  in  a  one  per  cent,  solution  of  acetic  acid.  This  brightens 
the  colour  and  fixes  the  carmine  in  the  nuclei,  and  also  differentiates  the  stained  from  the 
unstained  parts.     The  stain  thus  becomes  more  selective. 


PICROCARMINE,  OR   PICROCARMINATE  OF   AMMONIA. 

Picrocarmine  Solution. — Make  a  solution  of  carmine  by  rubbing  up  i  grm.  of  best  carmine 
with  10  c.c.  of  water,  and  adding  3  c.c.  of  liquor  ammoniae.  Add  this  solution  to  200  c.c.  of  a 
cold  saturated  solution  of  picric  acid.  Leave  the  mixture  exposed  to  the  air  until  it  evaporates 
to  one-third  of  its  bulk.  The  same  result  may  be  obtained  by  evaporating  it  slowly  in  a  hot 
chamber,  or  over  a  water-bath  at  a  low  temperature.  Filter  to  get  rid  of  the  precipitated 
carmine. 

This  substance  was  introduced  by  Ranvier,  and  is  one  of  the  most  valuable  dyes  one  can 
employ.  It  is  especially  used  for  '  double  staining,'  containing  as  it  does  two  pigments,  one 
yellow  and  the  other  red — eg.  the  perinuclear  part  of  a  frog's  coloured  blood-corpuscle  is 


xliv  INTRODUCTION. 

stained  yellow,  the  nucleus  red.  Connective  tissue  is  stained  red  and  elastic  tissue  yellow. 
It  has  this  further  advantage — that  though  sections  are  left  for  several  hours  in  the  solution, 
they  seldom  become  overstained.  It  is  not  necessary  to  remove  all  the  picrocarmine  from 
the  section  ;  in  fact,  it  is  advisable  to  leave  a  little  adhering  to  the  section,  for  after  being 
mounted  in  glycerine  or  Farrant's  solution,  the  trace  of  dye  left  is  gradually  absorbed  by  the 
section.  If  it  be  desired  to  retain  the  yellow  and  red  colours  it  is  desirable  not  to  wasli  the 
section  after  staining,  because  the  yellow  picric  acid  stain  is  readily  removed  thereby.  In 
some  cases  it  is  advisable  to  mount  the  section  in  glycerine,  containing  one  per  cent,  of 
formic  acid. 

All  picrocarmine  preparations  improve  by  keeping.  After  several  days  nuclei  become 
stained  that  at  first  were  unaffected  ;  so  that  sections  should  invariably  be  examined  several 
days  after  they  are  mounted. 

Picrocarmine  may  be  purchased  as  an  imperfectly  crystalline  body.  If  the  crystals  arc 
used,  make  a  one  per  cent,  solution. 

Purpurine. —  The  dye  obtained  from  madder  has  no  advantages  over  any  of  the  other 
methods.     It  can  be  dispensed  with. 

ANILINE   DYES. 

{a)  Magenta  Solution.— Dissolve  i  decigrm.  of  the  sulphate  or  nitrate  of  rosanilin  in 
lOO  c.c.  of  water,  and  add  lO  c.c.  of  rectified  spirit.  This  does  very  well  for  fresh  tissues. 
The  stain  is  not  permanent,  though  it  may  last  for  many  months.  For  blood-corpuscles  2 
decigrms.  are  dissolved  in  40  c.c.  of  a  mixture  of  equal  parts  of  water  and  glycerine. 

{U)  Aniline  Blue,  soluble  in  Water. — Make  a  one  per  cent,  solution  in  water.  This  is 
sometimes  used  for  fresh  tissues. 

(c)  Aniline  Blue-Black. —  Make  a  one  per  cent,  solution  in  water.  If  the  section  is  over- 
stained  the  excess  may  be  removed  by  steeping  it  in  a  two  per  cent,  solution  of  chloral 
hydrate.  For  the  nervous  system  an  alcoholic  solution  is  most  useful.  Dissolve  i  decigrm. 
in  4  c.c.  of  water,  add  to  this  lOO  c.c.  of  rectified  spirit  and  filter.  Preserve  in  a  stoppered 
bottle.  This  solution  stains  rapidly  and  gives  a  pleasant  slate-grey  colour,  which  fatigues 
the  eye  very  slightly.  It  is  specially  useful  for  nerve-centres.  Sections  may  be  mounted  in 
dammar  without  any  fear  of  removing  the  dye. 

(d)  Methyl-Aniline. — Make  a  strong  watery  solution  and  stain  the  tissue  deeply.  Wash 
away  the  superfluous  pigment  in  water  acidulated  with  acetic  acid.  The  acetic  acid  dissolves  out 
much  of  the  pigment,  and  the  washing  must  be  continued  till  the  proper  tint  is  obtained. 
The  sections  must  then  be  thoroughly  washed  in  water,  and  mounted  in  glycerine  or  Farrant's 
solution.     Dammar  is  unsuitable,  as  clove  oil  discharges  the  colour. 

This  is  a  most  valuable  dye,  for  in  contact  with  certain  tissues  it  gives  a  '  double-stain.'  It 
decomposes  into  two  colours — one  a  red-violet,  the  other  a  blue-violet — each  of  which  acts  on 
different  tissues.  It  is  useful  for  hyaline  cartilage  ;  the  red-violet  attaches  itself  to  the  matrix, 
and  the  blue-violet  to  the  corpuscles. 

It  is  in  the  pathological  change  known  as  amyloid  or  waxy  degeneration,  however,  where  it 
is  most  useful.  All  parts  of  an  organ — e.g.  the  liver — affected  with  amyloid  degeneration  are 
stained  red-violet,  whilst  the  neighbouring  unaffected  elements  are  coloured  blue-violet. 

For  fresh  tissues  it  also  forms  a  useful  dye,  and  in  the  form  of  a  one  per  cent,  watery 
solution  it  may  be  used  instead  of  magenta.  It  stains  certain  parts  of  a  beautiful  violet.  It  is 
very  useful  for  showing  the  corpuscles  in  connective  tissue,  or  the  nuclei  in  fresh  cells,  or  for 
mucous  tissue.      It  is  best  to  mount  a  tissue  in  a  saturated  watery  solution  of  potassic  acetate. 


INTRODUCTION.  xlv 

{/)  Iodine  Green. — A  saturated  watery  solution  may  be  made  and  diluted  as  it  is  required, 
or  a  one  per  cent,  solution  may  be  prepared.  It  stains  rapidly,  and  the  stain  cannot  be 
removed  by  washing.  It  is  one  of  the  most  useful  of  the  aniline  dyes,  especially  for  double- 
staining  (p.  xlvi).  It  stains  mucous  glands  green,  and  acts  similarly  on  unossified  cartilage- 
Preparations  stained  with  it  must  be  mounted  in  dammar.  Do  not  leave  the  stained  sections 
long  in  spirit,  which  partially  removes  the  dye. 

(/)  Rosein  is  soluble  in  spirit,  and  so  is  useful  for  double-staining  when  the  sections  are  to 
be  mounted  in  dammar.     See  retina.    Ordinary  methylated  spirit  does  quite  well  as  a  solvent 

Many  other  aniline  dyes  are  employed,  but  the  above  are  certainly  the  most  useful. 

{g)  Eosin,  though  not  an  aniline  dye,  may  for  convenience  be  taken  here.  It  yields  a 
rose-coloured  dichroic  fluid,  when  dissolved.  Dissolve  i  part  in  1,500  of  water.  Tissues 
become  stained  in  a  few  seconds.  After  staining,  place  them  in  water  slightly  acidulated  with 
acetic  acid  (one  per  cent.),  to  remove  the  excess  of  pigment  and  to  'fix'  the  remainder. 
Sections  may  be  examined  either  in  glycerine  or  in  dammar.  For  some  purposes,  as  for 
staining  the  haemoglobin  of  coloured  blood-corpuscles,  it  is  used  as  an  alcoholic  solution.  It 
is  very  useful  in  the  study  of  developing  coloured  blood-corpuscles. 


METALLIC   SOLUTIONS. 

{a)  Silver  Nitrate. — Dissolve  i  grm.  of  silver  nitrate  in  100  c.c.  of  distilled  water.  For  use 
dilute  this  to  g^  or  ^  per  cent,  solution.  Wash  the  tissue  to  be  stained  {ivhick  must  be  quite 
fresh)  in  distilled  water  to  remove  chlorides,  place  it  in  the  silver  solution  for  five  minutes,  or 
until  it  becomes  of  a  whitish  appearance,  then  remove  it,  wash  it  in  ordinary  water,  and 
expose  it  in  water,  or  alcohol  and  water,  to  diffuse  daylight  till  it  becomes  brown  in  colour. 

Silver  nitrate  solution  is  used  where  it  is  desired  to  bring  into  view  the  existence  of  epithelial 
cells — e.g.  on  serous  membrane,  lining  arteries,  lymphatics,  capillaries,  or  the  air-cells  of  the 
lung.  It  acts  upon  the  intercellular  substance,  or  cement,  and  when  the  tissue  is  exposed  to 
the  action  of  light,  it  is  precipitated  in  the  intercellular  substance  as  the  black  oxide,  in  fine 
black  lines,  and  thus  the  cells  are  mapped  one  from  another.  These  lines  are  known  as 
'  silver  lines.'  It  also  stains  the  intercellular  substance  of  cartilage  and  the  cornea.  The  prepara- 
tions may  be  preserved  in  glycerine  or  dammar,  and  ought  to  be  kept  in  a  dark  place,  as  they 
are  apt  to  spoil  when  long  exposed  to  light. 

{b)  Gold  Chloride. — Solutions  from  \  per  cent,  to  2  per  cent,  are  employed. 

1.  Place  the  perfectly  fresh  tissue  in  the  gold  solution  (^  per  cent.)  for  twenty  to  thirty 
minutes,  wash  it  in  water,  and  then  expose  to  daylight  in  water  slightly  acidulated  with 
acetic  acid  (2  or  3  drops  of  acetic  acid  to  i  oz.  of  water)  until  it  becomes  of  a  purplish  colour. 

2.  Or  the  following  method  may  be  employed,  especially  where  the  tissue  is  dense  and  it  is 
not  desirable  to  retain  the  epithelium.  Cut  a  fresh  lemon  in  two,  squeeze  out  the  juice  and  fil- 
ter it  through  muslin.  Place  the  perfectly  fresh  tissue — e.g.  cornea — in  the  juice  for  five  or  seven 
minutes,  wash  it  in  wafer  to  remove  the  juice,  and  steep  it  in  a  one  per  cent,  solution  of  gold  for 
twenty  minutes,  or  half  an  hour.  Wash  off  the  surplus  gold,  and  place  the  object  in  a  mixture 
of  one  part  of  formic  acid  to  four  of  water.  Put  the  bottle  in  a  dark  place.  After  twenty- 
four  hours  the  gold  is  completely  reduced.  All  the  epithelium,  however,  is  removed  by  the 
formic  acid.     This  method  is  the  best  for  demonstrating  the  terminations  of  nerves  in  muscle. 

Gold  is  especially  useful  for  staining  nerve-fibrils — e.g.  in  the  cornea — and  also  for  connec- 
tive tissue-corpuscles. 

Under  each  tissue  where  gold  is  useful,  special  directions  are  given  for  its  employment. 


xlvi  INTRODUCTION. 

(c)  Osmic  Acid. — Make  a  one  per  cent,  solution  which  can  be  diluted  as  required — to  i  or  i 
per  cent.  The  method  of  using  and  preserving  this  substance  has  already  been  referred  to 
under  Hardening  Solutions  (p.  xxxii).  It  is  useful  for  blackening  fat-cells,  and  for  fixing  the 
white  substance  of  Schwann  in  nerve-fibres,  though  it  has  many  other  important  applications. 


ON    DOUBLE-STAINING   OF    TISSUES. 

By  using  two  differently  coloured  solutions,  it  is  found  that  in  certain  tissues  one  part 
will  take  up  one  colour,  whilst  another  part  takes  up  another.  Some  glands  stain  of  a  green 
colour  with  iodine-green — eg:  the  mucous  glands  of  the  tongue — whilst  the  other  glands,  the 
serous  glands,  are  not  affected  by  it. 

This  method  is  of  the  greatest  value,  and  I  find  it  can  be  practised  very  successfully  by 
students  even  in  large  classes. 

riCROCARMINE. 

As  already  indicated,  this  dye  stains  certain  parts  yellow  and  other  parts  red,  hence  its 
value. 

PICROCARMINE  AND   LOGWOOD. 

Stain  the  sections  slightly  with  picrocarmine,  and  after  washing  them  in  water,  place 
them  in  dilute  logwood  solution,  in  which  they  must  not  remain  too  long.  They  soon  be- 
come lilac-coloured.  Wash  them  and  mount  them.  This  method  is  useful  for  the  mesentery 
of  the  newt  (p.  38),  for  developing  bone,  epiglottis,  &c. 

PICROCARMIME  AND   IODINE   GREEN. 

This  is  one  of  the  most  useful  combinations  I  know.  Stain  the  sections  in  picrocarmine, 
and  wash  them  well  in  water  slightly  acidulated  with  acetic  acid,  and  then  stain  them  in  a 
watery  solution  of  iodine-green  (p.  xlv),  taking  care  that  they  do  not  become  over-stained, 
which  can  easily  be  ascertained  by  washing  them  in  water.  If  a  section  of  the  posterior  third  of 
the  tongue  be  so  stained  (p.  61),  all  the  muscles  and  connective  tissue  are  red,  whilst  the  mucous 
glands  and  adenoid  tissue  are  green.  The  serous  glands  do  not  take  up  the  green  stain.  This 
combination  is,  therefore,  of  the  utmost  value  for  gland-tissue.  All  the  mucous  glands  and 
adenoid  tissue  of  the  intestinal  tract  become  green,  whilst  the  connective  tissue  is  red,  and  the 
non-striped  muscle  of  a  light  yellow  or  brown.  Most  exquisite  effects  are  produced  in  the 
cerebellum,  bone,  and  intestine  by  this  method.     The  sections  must  be  mounted  in  dammar. 

EOSIN   AND   LOGWOOD. 

Stain  the  sections  lightly  in  eosin  and  then  in  logwood.  This  does  very  well  for  the 
brain. 

GOLD   CHLORIDE  AND   SILVER   NITRATE. 

This  method  is  sometimes  employed  for  the  cornea,  which  is  first  impregnated  in  the  usual 
way  with  gold  chloride  and  then  with  silver  nitrate.  The  method  is  very  difficult  to  be  suc- 
cessful with  and  often  fails. 


INTRODUCTION.  xlvii 


TREBLE-STAINING. 

This  method  has  recently  been  practised  by  Dr.  Heneage  Gibbes.  The  process  is  tedious, 
and  not  so  satisfactory  as  double-staining.  A  combination  of  picrocarmine  and  iodine-green 
for  sections  of  the  small  intestine,  including  a  Peyer's  patch,  gives  an  excellent  and  most 
instructive  effect  (p.  69),  in  fact  a  perfect  treble  stain. 

Other  combinations,  as  picrocarmine,  eosin,  and  iodine-green,  have  been  tried. 

GOLD   CHLORIDE  AND   AN   ANILINE   DYE. 

This  process  is  alluded  to  in  connection  with  the  study  of  the  parts  in  the  tail  of  a  rat 
(p.  25).  The  tissue  is  first  treated  with  gold  chloride  and  then  decalcified,  and  the  sections 
are  stained  with  rosein,  and  then  with  iodine-green,  or  with  iodine-green  alone.  The  effects 
produced  on  the  developing  bone  especially  are  most  remarkable  and  striking.  This  method  is 
capable  of  further  extension. 


LIST    OF   STAINING    REAGENTS. 

Ordinary  staining  reagents. 

1.  Logwood  or  HEematoxylin.  3.  Picrocarmine. 

2.  Carmine.  4.  Eosin. 

A  niline  dyes  soluble  in  water. 

1.  Magenta,  or  Rosanilin.  3.  Aniline  blue-black. 

2.  Aniline  blue.  4.  Methyl-aniline. 

5.  Iodine  Green. 

Aniline  dye  soluble  in  spirit. 
Rosein. 

Metallic  Substances. 
I.  Silver  Nitrate.  2.  Gold  chloride.  3.  Osmic  Acid. 

Many  other  dyes,  as  iodine,  indigo-carmine,  &c.,  are  recommended,  but  the  above  arc  what 
the  author  has  found  to  be  really  useful. 


xlviu 


INTRODUCTION. 


METHODS  OF  MOUNTING  SECTIONS. 

Most  sections,  even  when  very  thin,  are  too  opaque  to  be  examined,  and  must  therefore  be 
placed  in  some  medium  or  mounting  fluid  which  will  render  them  more  transparent. 

MOUNTING    FLUIDS. 

1.  Glycerine. The  section  must  be  floated  on   to  a  slide  from  water,  and,  after  removing 

the  water,  a  small  drop  of  glycerine  is  applied.  The  drop  ought  to  be  of  just  such  a  size 
that  when  the  cover-glass  is  applied,  it  accurately  fills  the  space  between  the  cover  and  the 
slide.  Apply  a  cover-glass,  as  directed  (p.  xxiv).  A  little  practice  is  required  to  gauge 
the  size  of  the  drop  required.  If  too  much  glycerine  be  added,  the  excess  is  most  conveniently 
removed  by  placing  several  small  pointed  pieces  of  bibulous  paper  round  the  margin  of  the 
cover.  All  surplus  glycerine  must  be  carefully  removed,  else  the  cement  to  be  afterwards 
applied  will  not  adhere  to  the  glass.  Glycerine  has  this  great  disadvantage,  that  the  cover- 
glass  moves  so  easily  that  it  becomes  difficult  to  seal  up  the  preparation.  This  is  entirely  obviated 
by  mounting  sections  in  the  following  fluid,  which  has  the  advantage,  that  after  exposure  to  the 
air  for  three  or  four  days,  the  cover-glass  becomes  firmly  fixed  to  the  slide,  so  that  the  coat- 
incT  of  cement  can  be  applied  without  any  fear  of  displacing  either  the  cover  or  the  section. 

2.  Farranfs  Solution. — Take  equal  parts  of  glycerine  and  a  saturated  watery  solution  of 
arsenious  acid.  Add  gum  arable  in  large  quantity,  and  allow  the  mixture  to  stand  for  several 
weeks,  stirring  it  from  time  to  time,  until  it  ceases  to  dissolve  more  of  the  gum.  Filter  through 
paper.     This  takes  a  considerable  time,  but  one  is   rewarded   by  the  beautifully  clear  fluid 

obtained. 

The  above  fluids  are  suitable  for  mounting  sections — stained  or  unstained  — from  water. 
Unless  for  very  delicate  tissues,  such  as  isolated  epithelial  cells,  Farranfs  solution  is  in  all 
cases  to  be  preferred.  It  is  always  advisable,  when  wishing  to  study  the  minute  relations  of 
parts— except  those  of  the  nerve-centres— to  mount  sections  in  either  of  the  above  fluids. 

3.  Glycerine  Jelly. This  is  sometimes  employed  to  mount  softened  bone  and  tooth,  and 

some  urinary  deposits.  It  must  be  melted  in  a  water  bath  before  being  used.  As  it  sets  rapidly, 
the  cover-glass  must  be  applied  at  once  to  the  preparation.  It  is  far  better  to  buy  it  ready- 
made  than  to  take  the  trouble  of  making  it.  Rimington's  is  the  best :  it  contains,  in  addition 
to  gelatine  and  glycerine,  a  trace  of  carbolic  acid. 

4.  Dammar  Solution. — Mix  in  an  earthenware  vessel  2  oz.  of  mastic  and  2  oz.  of  dammar  with 
2  oz.  of  chloroform  and  2  oz.  of  turpentine.  Shake  or  stir  the  mixture  from  time  to  time  until  the 
substances  are  dissolved.  Filter  through  paper.  Formerly  Canada  balsam  was  employed,  but 
the  above  fluid  is  certainly  to  be  preferred.  If  Canada  balsam  be  thought  preferable,  it  may  be 
easily  prepared  by  heating  some  balsam  in  a  capsule  in  a  warm  chamber  at  65°  C.  for  twenty- 
four  hours,  and,  after  it  cools,  dissolving  the  dry  yellow  mass  in  as  much  benzole  as  will  give  a 
fluid  of  a  syrupy  consistence. 


INTRODUCTION. 


XllX 


Both  dammar  and  Canada  balsam  render  tissues  much  more  transparent  than  glycerine. 

All  traces  of  water  must  be  removed  from  the  sections  before  they  can  be  mounted  in  dammar 
or  Canada  balsam.  This  is  done  by  immersing  them  for  five  or  ten  minutes  in  absolute 
alcohol. 

METHOD   OF    MOUNTING    IN    DAMMAR. 

When  a  section  is  to  be  mounted  in  dammar  all  water  must  be  got  rid  of.  Place  the 
section  in  absolute  alcohol  for  five  or  ten  minutes,  until  all  the  water  is  removed.  Transfer  it 
for  the  same  period  to  oil  of  cloves  or  turpentine, — preferably  the  former,  which  expels  the 
alcohol,  and  renders  the  section  quite  transparent.  These  operations  can  all  be  accomplished  on 
a  slide  thus, — Float  out  the  section  on  to  the  slide  in  water  (p.  xxvi)  ;  remove  the  water  with 
blotting-paper,  add  absolute  alcohol  to  remove  the  remainder  of  the  water  ;  after  a  few  minutes 
soak  up  the  alcohol  with  blotting-paper,  and  with  a  stiff  brush  insinuate  a  drop  of  oil  of  cloves 
under  the  section.  The  clove  oil  gradually  rises  through  the  section,  expels  the  alcohol,  and 
renders  the  section  quite  transparent.  This  process  ought  to  be  obser\ed  with  a  low  power 
under  the  microscope.  Remove  the  surplus  clove  oil  with  blotting-paper,  add  a  drop  of  the 
dammar  mounting  fluid,  apply  a  cover,  and  the  preparation  is  finished. 


LABELLING  AND  PRESERVING   SECTIONS. 

After  mounting,  each  section   must  be  properly  labelled  (p.  xxvi),  and  \a.\d  flat  in  a  tray, 
and  preserved  from  dust. 


iMETHODS    OF    SEALING    UP   THE    PREPARATIONS    FOR 

PRESERVATION. 


Each  preparation  has  still  to  be  sealed.  This  is  done  by  placing  some  kind  of  cement  over 
and  round  the  margin  of  the  cover-glass.  After  several  days  the  specimens  mounted  in  dammar 
and  Farrant's  solution  will  be  ready  for  sealing  up.  This  is  best  done  by  means  of  a  turntable, 
which  consists  of  a  brass  disc,  at  least  three  inches  in  diameter,  and  rotating  on  a  pi\ot. 
The  upper  surface  of  the  disc  has  on  it  a  series  of  concentric  rings,  corresponding  to  the  different 
sizes  of  cover-glasses  used.  The  disc  is  rotated  by  means  of  the  forefinger  of  the  left  hand 
acting  on  a  small  milled  head  placed  underneath.  The  slide  is  placed  on  the  disc,  accurately 
centred  and  fi.xed  in  position  by  two  clips,  which  ought  to  be  made  of  bent  steel  spring 
in  preference  to  brass.  A  camel-hair  or 
goat-hair  brush  is  dipped  in  the  cement, 
and  whilst  the  disc  is  rotated  a  ring  of  the 


cement  is  run  round  and -over  the  margin 


of  the  cover.  This  ought  to  be  done 
twice  or  thrice,  until  a  sufficiently  strong 
ring  is  formed.  In  selecting  a  turntable, 
see  that  the  disc  is  heavy  and  wide  enough 
to  admit  the  largest  slide  under  the  clips. 
The  disc  must  be  heavy,  and  should  rotate 
steadily  and  easily ;  and  this  is  best  secured  by  having  the  pin  or  pivot  on  which  it  rotates  made 

f 


Fig.  22.    Modified  Turntable. 


1  INTRODUCTION. 

^pin-pointed',    i.e.  the  upper  half  of  the  pin   much  finer  than  the  lower  half.     When  square 
cover-glasses  are  used,  the  cement  is  simply  painted  on  with  a  brush. 

Fig.  22  shows  a  turntable  invented  by  Mr.  Dunning.  It  is  so  made  that  any  non-central 
position  may  be  obtained,  even  with  slides  two  inches  in  width,  thus  facilitating  the  appli- 
cation of  the  cement  to  any  slide  the  cover-glass  of  which  has  not  been  accurately  centred. 

I.   METHODS   FOR   TREPARATIONS   MOUNTED   IN   DAMMAR. 

It  is  not  necessary  to  apply  any  cement ;  but  the  preparation  looks  neater  if  a  ring  of  the 
ordinary  dammar  mounting  fluid  is  run  round  the  margin  of  the  cover-glass.  After  using 
the  brush  wash  it  in  benzole  to  remove  the  dammar. 

II.   FOR   PREPARATIONS   MOUNTED   IN    GLYCERINE   OR   FARRANT'S   SOLUTION. 

After  removing  all  the  surplus  mounting  fluid,  paint  a  ring  of  luarinc  ghte  ox  painter  s gold- 
size  round  the  cover-glass,  so  as  to  fix  it  down,  and  after  it  dries  (twenty-four  hours)  put  on  a 
tliick  ring  of  zinc-white  cement.  It  may  be  necessary  to  put  on  more  than  one  ring  of  zinc 
cement.  This  is  by  far  the  best  way  of  sealing  up  these  preparations.  When  the  zinc-white 
dries  it  forms  a  ring  as  hard  as  enamel.  The  brush  used  for  the  zinc-white  ought  to  be 
washed  at  once  in  benzole,  and  the  one  used  for  gold-size  in  turpentine,  and  the  one  for  glue 
in  water.     These  simple  precautions  ought  never  to  be  neglected. 

Various  other  substances,  as  asphalt  and  melted  paraffine  are  recommended  for  sealing  up 
preparations,  but  none  of  them  are  equal  to  the  above  described  method. 

The  jEsthetically  inclined  student  can  easily  tint  the  white  cement  by  adding  to  it  a  little 
solid  eosin  or  any  other  dye.  It  is,  however,  quite  unnecessary  to  spend  time  in  adorning  the 
outer  surface  of  preparations. 

FLUIDS    FOR    SEALING    UP    PREPARATIONS. 

1.  The  ordinary  dammar  solution  may  be  used. 

2.  Zinc-white  Cement. — (Gum  dammar,  8  oz.  ;  zinc  oxide,  i  oz.  ;  benzole,  8  oz.  Dissolve 
the  dammar  in  the  benzole  and  add  the  zinc  oxide,  and  then  strain  through  muslin.) 

3.  Marine  Glue  (Hollis'). 

4.  Painter's  Gold  Size. 

Number  3  or  4  is  applied  first,  and  when  it  becomes  dry  it  is  covered  with  a  ring  of  zinc- 
white  cement.  In  my  opinion  gold  size  is  preferable  to  marine  glue.  It  is  not  so  liable  to 
crack. 


METHODS    OF    INJECTING   THE    BLOOD-VESSELS. 

The  ordinary  method  of  hardening  tissues  in  chromic  acid  does  not  suffice  to  reveal  the 
arrangement  of  the  blood-vessels  of  a  part,  hence  we  must  have  recourse  to  filling  the  blood- 
or  lymph-vessels  artificially  with  a  coloured  mass.  The  coloured  fluids  at  present  employed 
are  of  two  kinds — one  kind  which  \s.  fluid,  the  other  which  is  solid  at  ordinary  temperatures. 
The  chief  medium  of  the  former  is  water  or  water  and  glycerine,  and  of  the  latter  gelatine. 


INTRODUCTION.  li 

INJECTION    FLUIDS. 

(a.)    injection-masses  pxuid  at  ordinary  temperatures. 

1.  Watery  Solution  of  Briicke's  soluble  Berlin-blue. — Dissolve  lo  grms.  of  Briicke's  soluble 
blue  in  500  c.c.  of  distilled  water. 

Briicke's  soluble  blue  can  be  purchased  in  the  market  ;  but  if  it  be  desired  to  make  it,  the 
following  is  the  formula.  The  process  is  rather  tedious,  {ci)  Dissolve  217  grms.  of  potassic 
ferrocyanide  in  i  litre  of  distilled  water,  {b)  Make  one  litre  of  a  10  per  cent,  solution  of  ferric 
chloride.  Add  to  each  of  these  solutions  {a  and  b)  two  litres  of  a  saturated  solution  of  sodic 
sulphate.  Then  add  the  ferric  chloride  to  the  ferrocyanide,  and  stir  the  fluids  whilst  they 
mix.  The  precipitate  is  filtered  through  a  conical  bag  of  new  flannel,  and  washed  with  a 
small  quantity  of  water.  The  first  washings  are  returned.  The  washing  is  repeated  for 
several  days,  until  the  solution  when  passed  through  filter  paper  is  of  a  deep  blue  colour.  It 
is  then  soluble.  The  precipitate  is  collected  from  the  flannel,  pressed  between  folds  of 
blotting  paper,  dried,  and  broken  into  small  pieces.  After  the  blood-vessels  of  an  organ  have 
been  filled  with  this  solution  it  is  placed  in  equal  parts  of  methylated  spirit  and  water,  to 
which  a  trace  of  hydrochloric  acid  is  added,  as  this  prevents  the  diffusion  of  the  blue  ;  or  the 
organ  may  be  hardened  in  Miiller's  fluid  or  picric  acid.  In  some  cases  it  is  advisable  to  heat 
the  injection-fluid  to  a  temperature  of  40°  C.  before  injecting  it  into  the  vessels.  A  little 
glycerine  added  to  the  mass  makes  it  flow  more  easily. 

2.  Richardson's  Blue. — {a)  Dissolve  10  grains  of  ferric  sulphate  in  10  oz.  of  water,  [b) 
Dissolve  32  grains  of  potassic  ferricyanide  in  i  oz.  of  water.  Mix  a  and  b,  and  then  add 
water  to  20  oz. 

(b.)    injection-masses  not  fluid  at  ordinary  temperatures. 

1.  Soluble  Blue  and  Gelatine. — Soak  50  grms.  of  the  best  gelatine  (Cox  or  Coignet)  in 
water  for  several  hours.  Pour  ofl"  the  water  which  is  not  absorbed  by  the  strongly  swollen-up 
gelatine,  and  melt  the  gelatine  over  a  water-bath.  The  gelatine  may  be  placed  in  an  ordinarj- 
jelly  can  or  tin  vessel,  and  this  placed  in  a  pan  of  water  on  the  fire,  if  a  water-bath  is  not 
at  hand.  A  two  per  cent,  watery  solution  of  Berlin-blue  is  prepared  (p.  li.),  and  heated  to 
the  same  temperature  as  the  gelatine.  Take  250  c.c.  of  this  fluid,  and  gradually  add  to  it  the 
solution  of  gelatine,  the  mixture  being  stirred  all  the  time.  Whilst  still  hot  the  mixture  is 
filtered  through  flannel,  and  then  kept  at  a  temperature  of  40°  C.  until  it  is  injected.  After 
the  injection  is  finished,  place  the  organ  in  alcohol  for  twenty-four  hours,  to  coagulate  the 
gelatine. 

2.  Carmine  and  Gelatine  Mass.— («)  Take  i  oz.  of  the  best  gelatine,  and  make  a  solution 
of  it  as  described  above.  Strain  while  hot  through  flannel,  and  make  up  the  solution  to  2  oz. 
{b)  Place  I  drachm  of  the  best  carmine  in  a  mortar,  to  it  add  i  drachm  of  liquor  ammonia;  and 
2  oz.  of  water,  and  leave  it  for  twelve  hours  (if  in  a  hurry,  place  the  carmine  solution  near  the 
fire  for  an  hour).  Filter,  and  add  about  80  minims  of  glacial  acetic  acid,  drop  by  drop,  stirring 
all  the  while,  until  the  ammonia  is  completely  neutralised.  As  the  odour  of  the  ammonia 
becomes  faint  the  acid  must  be  added  very  cautiously.  As  long  as  there  is  free  ammonia  the 
fluid  is  of  a  dull  red,  but  it  becomes  of  a  bright,  florid,  cherry  colour  the  moment  the  ammonia 
is  neutrali.sed.  Add  the  florid  red  carmine  fluid  (Ji)  to  the  2  oz.  of  gelatine  solution  {a),  and  keep 
at  a  temperature  of  40°  C.  until  the  mass  is  required. 

f  2 


lii 


INTRODUCTION. 


The  principle  to  be  attended  to  in  making  the  mass  is  this  :— The  carmine,  if  alkaline, 
would  diffuse  through  the  tissues  ;  if  acid,  it  would  be  deposited  in  fine  granules,  which 
block  up  the  capillaries— hence  the  necessity  for  having  a  neutral  fluid.  The  best  guides 
are  the  colour  and  odour  of  the  fluid.  The  injected  organ  must  be  placed  in  equal  parts  of 
methylated  spirit  and  water,  to  which  a  few  drops  of  acetic  or  hydrochloric  acid  (one  per 
cent.)  are  added,  to  prevent  the  carmine  from  diffusing  into  the  tissues. 

APPARATUS    EMPLOYED    FOR    INJECTING   THE    BLOOD-VESSELS. 

Injections  are  made  either  with  a  syringe,  or  with  .some  form  of  apparatus,  which  will  give 
a  constant  pressure.     Whatever  apparatus  be  employed  it   is  necessary   to  insert   a  cannula 

into  the  blood-vessel  by  which  the  injection 
will  flow,  e.g.  the  aorta.  The  cannulae  may 
be  made  of  brass,  such  as  are  ordinarily 
supplied  with  a  syringe,  or  they  may  be 
made  of  glass,  with  a  short  piece  of  india- 
rubber  attached  to  them.  The  advantage 
of  a  glass  cannula  is  that  one  can  see  when 
air-bubbles  are  present.  For  the  method 
of  making  these  cannula  see  fig.  23. 

The  cannula  ought  to  have  a  shoulder, 
over  which  the  thread  is  applied,  so  that 
when  tied  it  cannot  slip  out  of  the  vessel. 

Method  of  inserting  a  Cannula  into  a 
Blood-vessel.  —  Expose  the  artery  of  the 
part  to  be  injected,  clear  a  small  piece  of 
it  from  the  surrounding  tissue,  and  place 
a  ligature  around  it  by  means  of  an 
aneurism-needle,  or  a  narrow-pointed  pair  of  forceps.  With  sharp  scissors  make  an  oblique 
slit  in  its  wall,  and  insert  the  nozzle  of  the  cannula,  directing  it  towards  the  periphery.  Tie 
the  ligature  firmly  over  the  artery,  and  behind  the  shoulder  on  the  cannula. 


Fig.  23.    Cannul.e  for  Injecting.     Natural  Size. 

r'  f- 1-^  Glass  Cannula;  of  different  sizes;  a  and  b  show  the 
method  of  making  a  cannula  :  a  glass  tube  is  heated  in  a 
flame  and  drawn  out  lo  the  size  required,  as  at  a.  It  is 
then  heated  in  the  middle  and  slightly  drawn,  to  make  it 
thinner  there.  After  it  cools  it  is  cut  by  means  of  a  triangular 
file  at  /,  and  then  each  half  is  ground  down  obliquely  on  a 
hone.  The  sharp  edges  are  got  rid  of  by  heating  it  in  a  flame 
for  a  few  seconds.  A  piece  of  india-rubber  tubing  is  tied  to  the 
end  of  each  and  the  cannula  is  complete.  c\  A  metal  cannula ; 
cl,  A  steel  clamp  for  clamping  a  small  artery  (Schafer). 


HOW   TO   INJECT   BY   MEANS   OF   A   SYRINGE. 

The  syringe  is  made  of  brass,  and  is  provided  with  several  cannulae,  of  different  sizes,  and 
a  separate  piece  with  a  stopcock  to  connect  the  cannula  with  the  syringe.  Tie  the  cannula  in 
the  vessel,  as  directed  above,  and  fill  it  with  salt  solution,  by  means  of  a  pipette.  Fix  the 
stopcock-piece  into  the  cannula,  and  fill  it  in  the  same  way  with  salt  solution.  Then  fill 
the  syringe  with  the  injection-mass,  taking  care  not  to  suck  up  any  air.  All  air  ought  to  be 
expelled  from  the  syringe  by  holding  it  vertically,  nozzle  uppermost,  and  expelling  a  few  drops 
of  the  injection-mass.  The  syringe  ought  to  be  filled  and  emptied  several  times  before  doing 
this,  to  get  rid  of  air.  Insert  the  syringe  in  the  stopcock-piece,  and  proceed  slowly  with  the 
injection.  The  piston  must  be  rotated  very  slowly;  so  that  the  mass  is  driven  on  slowly  and 
steadily.  Half-an-hour  is  not  too  long  for  the  injection  of  a  rabbit.  One  judges  of  the  com- 
pleteness of  the  injection  by  observing  the  colouration  of  such  vascular  parts  as  the  tongue, 
lips,  toes,  or  ear.  After  the  injection  is  completed,  turn  the  stopcock  to  prevent  the  escape  of 
fluid,  remove  the  syringe,  and  place  the  part  injected  in  equal  parts  of  alcohol  and  water,  or 
any  of  the  other  fluids  described  at  p.  xxxii.     If  a  gelatine  mass  be  employed,  it  is  better  to 


INTRODUCTION. 


liii. 


inject  the  animal  after  it  has  been  killed  by  bleeding.  The  animal  or  part  to  be  injected  must 
be  kept  warm  by  immersing  it  in  water  at  40°  C.  Even  when  a  watery  injection  is  emploj-ed, 
it  is  as  well  to  warm  it  to  40°  C,  as  while  hot  it  causes  less  spasm  of  the  small  arteries  than  a 
cold  mass. 

The  art  of  injecting  can  only  be  acquired  by  practice.  The  part  to  be  injected  and  the 
mass  must  be  at  the  proper  temperature,  no  air  must  be  allowed  to  enter  the  blood-vessels, 
and  the  mass  itself  must  not  contain  any  particles  which  will  block  up  the  capillaries.  One  of 
the  great  difficulties  is  to  keep  the  pressure  constant  ;  this  may  in  part  be  arrived  at  by  fi.x- 
ing  a  tube  at  right  angles  to  the  side  of  the  stopcock-piece,  and  bringing  in  connection  with  it, 
by  means  of  a  caoutchouc  tube,  an  ordinary  mercurial  manometer,  viz.,  a  bent  tube  filled  with 
mercury.  A  pressure  of  two  to  five  inches  of  mercury  is  usually  sufficient  for  injecting  small 
animals. 

In  selecting  a  syringe  sec  that  you  get  one  that  is  capable  of  holding  from  three  to  six 
ounces.  The  barrel  ought  to  be  /c//^— six  or  seven  inches.  For  corrosive  (or  metallic)  sub- 
stances, a  glass  or  vulcanite  syringe  must  be  employed. 


INJECTING   APP.\RATUS   FOR   OBTAINING   A   CONSTANT   PRESSURE. 

Ludwig  was  the  first  to  introduce  this  arrangement.     Sufficient  pressure  may  be  obtained 
either  b\'  means  of  a  column  of  mercury  or  a  column  of  water. 


Fig.  24.    Ludwig's  Mercury  Pressure-api'aratu.s. 

a.  Reservoir  bottle  for  mercury;  b,  Pressure  bottle  inio  which  the  mercury  of  a  flows;  c.  Bottle  containing  the 
injection-mass,  connected  with  b  by  one  india-rubber  tube,  and  by  another  with  the  cannula  f,  and  by  a 
glass  tube  with  a  small  manometer ;  d.  Handle  of  screw  for  raising  the  wooden  shelf  on  which  a  rests  ;  e, 
A  screw  clip,  and  g,  a  spring  clip. 

Ludwig's  Mercury  Pressure-apparatus  (fig.  24). — The  apparatus  consists  of  two  Wolff's 
bottles,  one  of  which,  a,  contains  a  reserve  of  mercury,  and  ^>,  into  which,  owing  to  the  bottles 
being  at  different  levels,  the  mercury  flows.     This  compresses  the  air  in  d,  and  so  acts  on  the 


liv 


INTRODUCTION. 


injection-fluid  contained  in  the  bottle  c,  to  which  a  mercurial  manometer  is  attached.  From 
this  an  exit  caoutchouc  tube  passes,  which  is  attached  to  the  cannula  fixed  in  the  artery.  The 
pressure-bottle  a  rests  on  a  wooden  support,  which  is  raised  by  means  of  a  screw,  and  in  this 
way  sufficient  pressure  is  obtained.  In  injecting  with  any  pressure-apparatus,  begin  with  a 
pressure  of  one  inch  of  mercury,  and  slowly  increase  it  to  four  inches.  Of  course  if  a  gelatine 
mass  be  employed  the  bottle  c  must  be  kept  warm,  by  placing  it  in  warm  water  to  keep  the 
gelatine  in  a  fluid  condition. 

Before  beginning  the  injection,  the  cannula  and  the  exit-tube  from  c  must,  of  course,  be 
filled  with  the  injection-mass,  to  avoid  the  entrance  of  air.  This  apparatus  is  expensive,  and 
therefore  an  apparatus  that  one  can  make  for  himself  is  to  be  preferred  ;  such  as  the  following 
simple  mercurial  injection-apparatus  (fig.  25).      It  works  on  exactly  the  same  principle,  and 


Fig.  25.    Simple  Mercurial  Injection-apparatus. 


fig.  25  explains  itself.  One  of  the  bulbs  (c),  containing  the  mercury,  is  stationary,  whilst 
the  other  (a)  rests  in  a  wooden  framework,  or  shelf,  which  moves  freely  in  a  slot  (d). 

The  part  of  this  movable  shelf  which  is  fi.xed  in  the  slot  is  so  made  that  it  works  very 
freely,  and  when  it  is  loaded  at  the  free  end  of  the  horizontal  arm,  it  becomes  self-fixing  at 
any  height.  The  pressure  is  very  easily  regulated  by  raising  or  depressing  the  movable  shelf 
(a).  By  means  of  a  divided,  or  Y-shaped  tube,  two  injection-fluids  (b,  r)  may  be  used  simul- 
taneously.    This  arrangement,  I  find,  works  admirably. 

Simple  Water  Pressure-apparatus. — Mercury  is  not  always  to  be  obtained,  hence  it  is 
sometimes  convenient  to  employ  water-pressure.  Satisfactory  results  are  not  obtained  by 
connecting  a  water-tap  to  a  system  of  pressure-bottles.  The  following  arrangement  (fig.  26) 
is  simple,  cheap,  easily  made,  and  effective.  It  consists  of  a  bottle,  C,  which  holds  the  injection 
mass,  a  much  larger  glass  bottle,  B,   with  a  manometer  (m)  attached  ;  and  communicating 


I  NT  ROD  UCTION. 


Iv 


with  this  is  a  cyhiider  of  tin  (a),  which   is  filled  with  water,  and  is  raised  or  lowered  by 
means  of  a  cord  which  passes  over  a  pulley  (p)  fixed  to  the  ceiling  of  the  room. 


INJECTION   OF   THE   BLOOD-VESSELS   OF  AN   ENTIRE  ANIMAL. 


Having  determined  which  injection-mass  is  to  be  used,  prepare  the  cannula;.  The  animal 
—a  rabbit  or  guinea-pig— whose  blood-vessels  are  to  be  injected,  is  killed  by  chloroform.  As 
soon  as  it  ceases  to  breathe,  the  thorax  is  opened  by  making  an  incision  along  the  middle  of 
the  sternum,  so  as  to  expose  the  heart.  Pull  the  heart  forward,  and  open  the  pericardium. 
Snip  through  the  wall  of  the  right  auricle,  and 
allow  as  much  blood  to  flow  out  as  possible ;  the 
animal  may  even  be  suspended  by  the  hind  limbs 
to  expel  as  much  blood  as  possible.  Expose  the 
aorta,  and  tie  into  it  a  glass  cannula  (fig.  23,  c) 
as  directed  at  p.  lii.  The  cannula  is  now  filled 
with  salt  solution,  and  is  connected  either  with  a 
syringe  or  one  of  the  constant-pressure  arrange- 
ments described  above.  Take  care  that  there 
is  no  air  either  in  the  cannula  or  in  the  india- 
rubber  tube  coming  from  the  injection-bottle.  At 
first  a  pressure  of  two  inches  of  mercury  is  suffi- 
cient, but  gradually  the  pressure  may  be  raised  to 
four  inches.  As  the  injection  is  forced  into  the 
aorta,  and  gradually  fills  the  blood-vessels,  blood 
and  then  blood  and  injection-mass  flow  out  of 
the  right  auricle.  Continue  the  injection  until  the 
injection  alone  flows  from  the  hole  in  the  right 
auricle.  It  is  then  closed  by  a  broad  ligature,  and 
the  injection  continued  until  the  vascular  system  is 
filled  with  the  injection,  which  may  be  judged  of 
by  the  colour  of  transparent  parts,  as  the  ears,  lips, 
&c.  After  the  injection  is  complete,  ligature  the 
vessels  at  the  base  of  the  heart,  or  a  glass  stopper 
ma)^  be  put  into  the  caoutchouc  tube  attached  to 
the  glass  cannula.  The  after-treatment  of  the  in- 
jected animal  is  described  at  p.  lii. 

If  an  animal  be  allowed  to  cool  after  death,  it 
must  be  thoroughly  warmed  before  using  a  gelatine 
mass.  This  is  done  by  placing  it  for  an  hour  in 
w'arm  water  not  above  40°  C. 


Fig.  26. 


Injection  Apparatus  with  Water 
Pressure. 


A,  Vessel  filled  with  water;   B,  Bottle  for  compressed 
air ;  c,  Bottle  for  injection  mass ;  M,  Manometer. 


INJECTION    OF   THE   LYMPHATICS. 

Puncture  Method. — This   is  very  easily  done  by  the  '  einstich  '  or  '  puncture  '  method  of 
Ludwig.     A  small  subcutaneous  (Pravaz)  syringe  (fig.  27,  p.  22)  is  filled  with  a  two  per  cent. 


Ivi  INTRODUCTION. 

solution  of  Briicke's  blue  (p.  li),  and  the  nozzle  is  thrust  into  the  pad  of  a  dog's  or  cat's  foot, 
and  the  fluid  injected  into  the  pad,  which  is  then  compressed,  and  the  limb  stroked  from  below 
upwards.  The  coloured  mass  finds  its  way  from  the  connective-tissue  spaces  into  the 
lymphatic  vessels,  and  runs  along  them  very  earily.  In  this  way  one  can  easily  inject  the 
glands  of  the  leg  and  abdomen,  and  even  the  thoracic  duct.  The  lymphatics  of  the  testis  are 
injected  in  the  same  way.  It  is  easy  to  demonstrate  the  lymphatics  in  the  intestinal  wall  and 
in  the  liver  by  the  same  method.  Instead  of  a  syringe,  one  of  the  constant-pressure  arrange- 
ments may  be  used. 


PRACTICAL     HISTOLOGY. 


o»{o 


BLOOD. 

When  examined  microscopically  blood  is  seen  to  contain  corpuscles  floating  in  a  fluid — the 
plasma  or  liquor  sanguinis.  The  corpuscles  are  of  two  kinds,  (i)  coloured  and  (2)  colour- 
less, and  in  addition  (3)  minute  granules  are  to  be  seen.  Besides  these,  there  fall  to  be 
examined  (4)  the  fibrin  and  (5)  the  colouring  matter  of  the  coloured  corpuscles  and  (6) 
various  pigments  derived  therefrom. 

(A.)     BLOOD   OF    A    FROG    OR    NEWT. 

PREPARATION. — Give  a  frog  or  newt  a  smart  blow  on  the  head.  If  a  newt  be  taken, 
carefully  wipe  its  tail  dry  by  means  of  a  cloth  and  snip  off  the  point  of  the  tail.  Then 
squeeze  out  a  drop  of  blood  from  the  stump  on  to  a  glass  slide,  and  at  once  apply  a  cover- 
glass.  If  a  frog  be  used  a  toe  should  be  cut  off,  when  a  drop  of  blood  can  easily  be  squeezed 
out  of  the  stump.  In  cither  case  the  blood  thus  obtained  will  be  mixed  with  a  considerable 
quantity  of  lymph.  If  a  drop  of  blood  without  any  admixture  of  lymph  is  required,  the  heart 
of  the  animal  must  be  exposed,  and  when  this  is  done  the  auricle  should  be  snipped  through 
and  a  drop  of  blood  allowed  to  fall  on  the  glass  slide. 

EXAMINATION. —  i.  Observe  the  coloured  corpuscles  (H).  They  are  seen  to  be  very 
numerous,  of  a  yellowish  straw  colour,  uniform  in  size  and  elliptical  in  shape  when  seen  on  the 
flat  (PI.  I.,  Fig.  i).*  A  few  of  them  may  also  be  seen  edgewise,  and  they  then  appear  narrow, 
somewhat  pointed  at  the  ends,  and  slightly  thicker  in  the  middle  (PI.  I.,  Fig.  2).  Sometimes 
one  corpuscle  lies  above  another,  when  the  outline  of  the  undermost  one  may  be  distinctly 
seen  through  the  one  lying  above  it ;  it  is  thus  seen  that  each  corpuscle  is  transparent. 
At  first  each  corpuscle  may  seem  homogeneous,  but  in  a  very  short  time  an  elliptical  slightly 
granular  body  shows  itself  in  the  centre  of  each  as  seen  on  the  flat.  This  body,  which  is 
somewhat  lighter  in  colour  than  the  rest  of  the  corpuscle,  is  called  the  )U(deiis  (PL  I.,  Fig.  i). 

EFFECTS  OF  REAGENTS  on  the  coloured  corpuscles. 

{a)  One  per  cent.  Hydro cMoric  Acid. — Squeeze  out  a  fresh  drop  of  blood  on  a  clean  slide, 
quickly  apply  a  cover-glass,  and  then  place  a  drop  of  a  one  per  cent,  dilution  of  hydrochloric 
acid  at  one  edge  of  the  cover-glass  (taking  care  that  none  of  the  acid  gets  on  the  top  of  the  cover- 

»  Where  a  simple  reference  is  made  to  a  plate  and  figure  (as  PI.  I.,  Fig.  i),  it  is  intended  that  the  student  shall  insert 
by  delineation  and  colouring  such  appearances  and  changes  as  are  indicated  in  the  text  as  the  result  of  reagents  or  otherwise. 


2  PRACTICAL   HISTOLOGY. 

fjlass).  Place  a  small  piece  of  blotting-paper  on  the  slide  at  the  edge  of  the  cover-glass  opposite 
to  the  spot  where  the  drop  of  acid  was  placed,  which  will  have  the  effect  of  drawing  the  acid 
under  the  cover  so  as  to  '  irrigate '  the  blood-corpuscles  with  the  reagent.  Steadily  observe 
one  or  two  corpuscles  ;  they  will  gradually  enlarge,  become  globular  and  all  of  a  sudden  swell 
up  and  become  distinctly  spherical,  and  then  as  suddenly  collapse  and  '  shoot '  or  discharge  their 
hasmoglobin.  The  nucleus  comes  clearly  into  view  during  the  process.  This  process  ought  to 
be  diligently  looked  for,  because  the  phenomenon  is  so  striking.  It  is  as  if  the  corpuscle  burst 
and  discharged  something.  After  the  collapse,  the  residue  of  the  stroma  adheres  to  the  nucleus. 
In  other  cases  the  corpuscles  do  not  collapse,  but  remain  globular,  clear,  and  transparent.  The 
peri-nuclear  part  becomes  decolourised,  and  appears  to  be  limited  externall}^  by  a  membrane 
from  which  fine  shreds — the  residue  of  the  stroma — stretch  to  the  nucleus,  which  is  now  clearly 
brought  into  view,  and  has  assumed  a  distinctly  granular  appearance,  due  to  the  existence  of 
a  fibrillar  network  in  its  interior.  If  the  acid  used  be  too  strong  this  action  may  be  missed, 
and  the  outline  of  the  corpuscles  becomes  very  faint,  and  the  shrivelled  nucleus  may  appear 
excentrically,  or  may  even  escape  altogether  outr-ide  the  corpuscle.  These  phenomena  are 
best  observed  with  frog's  blood. 

(/;)  Acetic  Acid. — Shows  the  same  phenomena,  though  in  a  less  degree.  The  corpuscles 
swell  up  and  become  clear,  so  that  their  nucleus  becomes  very  obvious,  and  the  nucleus  not 
unfrequently  passes  out  of  the  corpuscles.  It  is  to  be  noted  that  all  the  corpuscles  are  not  so 
affected,  which  is  perhaps  due  to  difference  in  the  chemical  constitution  of  the  corpuscles.  In 
many  corpuscles  distinct  vacuoles  are  formed  in  the  peri-nuclear  part.  The  coloured  pig- 
ment may  be  seen  gradually  to  leave  the  wall  of  the  corpuscle,  and  become  heaped  up  round 
the  nucleus.  The  fluid  in  which  the  corpuscles  float  becomes  slightly  coloured  by  a  diffusion 
of  the  pigment  of  the  blood-corpuscle  (haemoglobin),  and  sometimes  the  nucleus  takes  up 
some  of  this  pigment  and  becomes  of  a  distinctly  yellow  colour  (PI.  I.,  Fig.  3). 

(c)  Water. — Squeeze  out  another  drop  of  blood  on  to  a  slide,  cover,  and  irrigate  with  water 
in  the  manner  just  described.  The  corpuscles  and  their  nuclei  both  swell  up  and  become 
globular,  owing  to  endosmosis,  and  at  the  same  time  the  haemoglobin  diffuses  into  the  sur- 
rounding fluid  (PI.  I.,  Fig.  6). 

{d)  Syrup. — Squeeze  out  a  fresh  drop  of  blood,  add  a  drop  of  strong  syrup  of  white  cane 
sugar,  and  mix  the  two  drops  thoroughly  with  the  point  of  a  needle,  and  apply  a  cover-glass. 
Owing  to  the  greater  specific  gravity  of  the  syrup  the  corpuscles  lose  fluid,  and,  on  examina- 
tion some  of  them  present  a  shrivelled  appearance,  and  resemble  thin  misshapen  biscuits, 
while  others  are  but  slightly  affected.  Here  and  there  a  reddish  tinge  is  seen  in  a  corpuscle 
(Pi.  I.,  Fig.  4). 

{e)  Dilute  Alcohol. — One  part  of  rectified  spirit  to  two  of  water,  applied  in  the  same  way 
as  acetic  acid,  decolourises  the  corpuscles,  causes  them  to  swell,  and  brings  a  nucleolus  into 
view  (usually  placed  towards  one  end)  in  each  nucleus  (Ranvier,  Stirling). 

(/)  Magenta. — Mix  a  drop  of  blood  with  a  drop  of  a  solution  of  sulphate  or  nitrate  of 
rosaniline  (p.  xliv),  cover  and  examine.  The  nuclei  of  the  coloured  corpuscles  become  stained 
of  a  deep  red  colour,  the  peri-nuclear  part  remaining  unaffected,  though  of  course  the  magenta 
has  to  traverse  it  to  reach  the  nucleus.  All  the  corpuscles  are  not,  however,  equally  affected. 
In  many  the  indications  of  the  existence  of  an  envelope  with  one  or  more  thickenings  in  it 
are  observable.  This  last  appearance  is  even  better  seen  if  the  magenta  be  added  after  the 
blood  has  been  irrigated  with  dilute  alcohol. 

[g)  Tannic  Acid. — Mix  a  drop  of  the  blood  with  a  drop  of  a  freshly  prepared  two  per  cent, 
aqueous  solution  of  tannic   acid   (made  with  hot   water   and   allowed   to  cool)  ;    cover   and 


BLOOD.  3 

examine.  The  tannic  acid  acts  on  the  serum-albumen  of  the  plasma  and  forms  a  finely  granu- 
lar precipitate  which  somewhat  obscures  the  field,  but  it  also  acts  (like  some  other  acids)  on 
the  corpuscles,  causing  them  to  become  globular  and  the  colouring  matter  to  separate  from  the 
stroma.  The  separated  colouring  matter  tends  to  pass  out  of  the  corpuscle,  but  as  it  does 
so  it  becomes  coagulated  by  the  acid,  and  remains  attached  to  the  corpuscle  in  the  form 
of  one  or  more  granular-looking  buds  (PI.  L,  Fig.  5). 

None  of  the  preparations  of  the  blood  of  the  frog  or  newt  hitherto  described  can  be 
permanently  preserved  ;  the  two  following  preparations  may,  however,  be  kept  as  permanent 
slides. 

(Ji)  Picric  Acid  and  Picrocarmine. — Place  a  drop  of  blood  on  a  slide,  and  add  a  drop  of  a 
saturated  solution  of  picric  acid,  put  the  slide  aside  and  allow  it  to  remain  for  five  minutes,  and 
at  the  end  of  that  time,  when  the  acid  has  '  fixed  '  the  corpuscles  (that  is,  has  coagulated  their 
contents),  the  excess  of  acid  should  be  removed  by  means  of  a  narrow  slip  of  blotting-paper. 
A  drop  of  a  solution  of  picrocarmine  (p.  xliii)  should  now  be  added,  and  a  trace  of  glycerine 
to  prevent  evaporation,  and  the  preparation  set  aside  for  an  hour.  At  the  end  of  that  time, 
remove  the  picrocarmine  solution  by  means  of  a  slip  of  blotting-paper,  and  add  a  drop  of 
Farrant's  solution  or  glycerine  (p.  xlviii)  and  apply  a  cover.  The  preparation  may  then  be  ex- 
amined, when  the  peri-nuclear  part  of  some  of  the  corpuscles  will  be  seen  to  be  highly  granular 
and  of  a  deep  yellow  colour  ;  while  the  nucleus  is  stained  red.  In  some  of  the  corpuscles 
there  may  also  be  seen  delicate  yellow-coloured  threads,  extending  from  the  nucleus  to  the 
envelopes.     In  others  the  peri-nuclear  part  remains  uniformly  homogeneous. 

(«')  Osmic  Acid  and  Picrocarmine. — Mix  a  drop  of  blood  with  a  drop  of  a  one  per  cent, 
aqueous  solution  of  osmic  acid  (p.  xlvi),  and  allow  the  slide  to  stand.  This  '  fixes  '  the  corpuscles 
without  altering  their  shape.  At  the  end  of  five  minutes  remove  the  excess  of  osmic  acid  with 
blotting-paper,  add  a  drop  of  solution  of  picrocarmine  and  a  trace  of  glycerine  to  prevent  evapo- 
ration, and  set  aside  for  three  or  four  hours  (or  even  longer,  as  no  over-staining  takes  place). 
At  the  end  of  this  time,  the  preparation  is  treated  as  iji)  and  examined.  The  nucleus  will 
now  be  found  to  be  stained  red,  and  the  peri-nuclear  part  homogeneous  and  yellow.  If 
a  drop  of  blood  (taken  from  a  frog  which  has  been  kept  through  the  winter)  be  prepared  by 
this   method,  some   of  the   corpuscles  may  show  '  vacuoles '  in  the  peri-nuclear  part.     (PI.  I., 

Fig.  7)- 

(j)  Osmic  Acid  and  Logwood. — A  similar  preparation  may  be  made  by  using  a  solution  of 
logwood  (p.  xlii)  as  the  staining  agent  instead  of  picrocarmine,  with  the  advantage  that  the 
staining  of  the  nucleus  takes  place  much  more  rapidly — in  about  five  minutes. 

ik)  Ammonium  Chromate  and  Picrocarmine. — Mix  a  few  drops  of  blood  with  a  small  quantity 
of  a  five  per  cent,  solution  of  ammonium  chromate  in  a  small  glass  vessel,  and  leave  the  mix- 
ture for  twenty-four  hours,  taking  care  to  prevent  evaporation.  Then  pour  off  the  chro- 
mate solution  and  substitute  picrocarmine  solution  for  another  twenty-four  hours.  Mount  a 
drop  of  the  deposit  in  glycerine  and  examine  it.  The  effects  of  this  substance  are  most 
remarkable.  Some  corpuscles  retain  their  shape  though  the  nucleus  enlarges,  and  shows  an 
intra-nuckar  plexus  of  fibrils.  In  others  the  haemoglobin  is  arranged  in  groups  around  the 
enlarged  nucleus  ;  while  others  have  their  wall  partially  dissolved  on  one  side,  with  part  of 
the  cell-contents  extruded  through  it. 

The  preparations  ij.)  and  (y)  show  colourless  corpuscles  with  their  nuclei  stained. 

2.  Observe  the  Colourless  Corpuscles  (H). — Prepare  a  drop  of  blood  as  for  the  examination 
of  the  coloured  corpuscles.     On  careful  examination  there  may  be  seen,  scattered  amongst 

u  2 


4  PRACTICAL   HISTOLOGY. 

the  coloured  corpuscles,  colourless,  irregular,  granular  bodies.  They  arc  few  in  number 
compared  with  the  red,  and  by  careful  searching  three  distinct  varieties  may  be  made  out 
as  follows  : — 

(a)  Finely  granular  Corpuscles.— These  have  no  definite  shape,  are  colourless,  and  have  a 
faint,  finely  granular  appearance.  They  are  usually  slightly  smaller  than  the  coloured 
corpuscles,  and  irregular  in  outline,  with  fine  projecting  processes.  Careful  observation  may 
discover  nuclei,  which  arc  irregular  in  shape,  and  sometimes  subdivided. 

(y8)  Coarsely  granular  Corpuscles. — These  also  are  indefinite  in  shape  and  colourless,  but 
contain  a  large  number  of  highly  refracted  granules,  usually  lying  at  one  side  of  the  corpuscle, 
whilst  the  subdivided  nucleus  occupies  the  other  side.  They  ma)'  be  either  larger  or  smaller 
than  (a)  and  their  processes  are  usually  blunt  and  rounded. 

(7).  The  third  variety  consists  of  small  nucleated  masses  of  protoplasm  about  the  size  of  the 
nucleus  of  a  coloured  corpuscle  ;  and  as  these  latter  sometimes  escape  from  the  corpuscles,  care 
must  be  taken  not  to  confound  the  one  with  the  other. 

Amoeboid  movements  of  Colourless  Corpuscles. — The  finely  granular  corpuscles  (a)  may  be 
seen  to  change  their  shape  and  place  (in  the  newt  even  at  the  ordinary  temperature  of  the  air) 
in  the  same  manner  as  an  amceba  does,  hence  the  term  amoeboid  ;  and  the  coarsely  granular 
corpuscles  (/3)  behave  in  a  similar  manner.  In  these  latter,  the  granules,  being  more  distinct, 
may  be  seen  as  it  were  to  rush  from  one  part  of  the  corpuscle  to  another.  In  this  change  the 
granules  are  passive,  they  arc  carried  by  the  protoplasm.  The  small  nucleated  masses  ^7) 
undergo  similar  changes  of  site  and  form,  but,  on  account  of  their  smallness,  these  phenomena 
are  not  so  easily  observable.  The  best  way  to  notice  these  changes  is  to  make  drawings  from 
time  to  time- — say  every  two  minutes — of  a  colourless  corpuscle.  The  change  of  form  is  some- 
times so  great  that  a  corpuscle  may  actually  be  seen  to  divide  into  two.  At  other  times  a 
transparent  spot  --a  vacuole — which  contains  fluid  may  be  seen  in  some  corpuscles.  All  these 
changes  take  place  more  rapidly  when  the  slide  is  warmed  on  a  hot  stage  (p.  xxviii)  ;  and 
when  this  is  done,  it  is  necessary,  in  order  to  prevent  evaporation,  to  surround  the  cover 
with  a  little  oil  or,  better,  with  a  ring  of  melted  paraffin.  {Sketch  the  changes  of  shape  in 
PI.  I.,  Fig.  8). 

Feeding  Colourless  Blood-corpuscles. — That  the  colourless  corpuscles  can  take  small  par- 
ticles into  their  interior  is  easily  demonstrated,  thus  :  all  that  is  required  is  to  rub  up  a  little 
china  ink  or  vermilion  in  a  three  quarter  per  cent,  salt  solution,  and  to  mix  a  drop  of  this 
with  the  blood  and  examine  the  preparation  from  time  to  time  on  a  hot  stage,  as  indicated 
above.  The  preparation  requires  to  be  sealed  up  with  oil  to  prevent  evaporation.  For  newt's 
blood  30°  C.  is  sufficient. 

The  EFFECTS  OF  REAGENTS  on  the  colourless  corpuscles  should  next  be  observed. 

{a)  Water. — When  the  blood  is  irrigated  with  water,  the  protoplasm  of  the  white  corpuscles 
swells  up,  becomes  quite  clear  and  transparent,  and  the  nuclei,  usually  two  or  three  in  number, 
come  clearly  into  view.  Sometimes  also  when  so  treated  the  granules  may  be  seen  to  exhibit 
Brovvnian  movement. 

{b)  Acetic  Acid. — When  blood  is  irrigated  with  a  one  per  cent,  dilution  of  acetic  acid,  the 
protoplasm  becomes  clear  and  transparent,  and  a  bi-  or  tri-partite  nucleus  is  distinctly  seen. 
{Indicate  this  effect  in  one  of  the  corpuscles  sketched  in  PI.  L,  Fig.  8.) 

{c)  Magenta. — When  the  blood  is  irrigated  with  magenta  solution  (p.  xliv),  the  protoplasm 
takes  on  a  faint  red,  the  nuclei  a  deep  red  colour.  {Indicate  the  effects  in  one  of  the  corpuscles 
sketched  in  PI.  I.,  Fig.  8.) 


Pi  ATE  I.  Blood 


■^^nt«l:T1.  Bros  lilh 


BLOOD.  5 

Note. — The  osmic  acid  and  picrocarmine,  and  osmic  acid  and  logwood  preparations  of 
blood  are  the  only  permanent  preparations  which  show  colourless  corpuscles.  A  careful  search 
is  sure  to  reveal  several  corpuscles  with  their  nuclei  stained. 

In  fresh  blood  an  eighth-of-an-inch  lens  shows  the  intra-nuclear  plexus  of  fibrils  in  the 
colourless  corpuscles. 

(B.)     HUMAN    BLOOD. 

PREPARATION. — Wrap  a  handkerchief  tightly  round  a  finger,  beginning  at  the  base  and 
proceeding  towards  the  point,  which  will  cause  the  point  of  the  finger  to  become  congested  ; 
then  prick  the  skin  at  the  root  of  the  nail  with  a  clean  sewing  needle  and  bring  a  slide  in  con- 
tact with  the  drop  of  blood  which  exudes.     Apply  a  cover  and  examine. 

EXAMINATION. —  i.  Observe  the  coloured  corpuscles  (H).  Af  fir.st  the  coloured  blood- 
corpuscles  will  be  seen  adhering  by  their  flat  surfaces  so  as  to  form  rouleaux.  These  piles 
may  form  a  kind  of  network.  [Indicate  these  rouleaux  in  PI.  I.,  Fig.  9.)  When  so  disposed 
it  is  of  course  their  edges  only  that  are  seen  ;  but  if  the  cover  be  gently  moved  with  a 
needle-point,  the  rouleaux  break  up,  and  then  the  individual  corpuscles  are  seen  both  on 
the  flat  and  on  edge.  On  the  flat  they  appear  as  bi-concave,  circular  discs  ;  while  edge- 
wise they  appear  somewhat  dumbbell-shaped  (PL  I.,  Fig.  10).  On  carefully  focussing  a 
corpuscle  as  seen  on  the  flat,  a  dark  part  at  first  appears  in  the  centre,  the  margin  being 
light  (PI.  I.,  Fig.  11);  but  on  altering  the  fine  adjustment  the  centre  becomes  light,  and 
the  circumference  dark  (PI.  I.,  Fig.  11).  These  changes  in  appearance  are  due  to  the  fact 
that  the  corpuscle  is  a  bi-concave  disc.  It  is  not  due  to  the  presence  of  a  nucleus,  for 
these  corpuscles  are  non-nucleated  and  homogeneous  throughout.  It  is  well  to  become 
familiar  with  the  size  of  a  corpuscle  seen  under  a  power  of  300  diameters,  for  a  corpuscle  so 
magnified  serves  as  a  useful  standard  of  comparison  for  other  objects.  The  diameter  of  a 
corpuscle  is  ^^rVo  inch.  It  is  generally  stated  that  the  coloured  corpuscles  are  all  of  the 
same  size,  but  this  is  not  strictly  correct,  for  a  careful  examination  shows  that  there  is  a 
slight  variation  from  this  standard,  and  this  variation  is  greater  in  some  individuals  than  in 
others,  and  becomes  specially  marked  in  some  diseases. 

EFFECTS  OF  REAGENTS,  [a)  Acetic  Acid.— When  irrigated  with  one  per  cent,  dilution 
of  acetic  acid,  the  coloured  corpuscles  become  globular  and  lose  their  colouring  matter,  which 
diffuses  into  the  surrounding  fluid,  so  that  it  is  difficult  to  distinguish  the  outline  of  the  indi- 
vidual corpuscles.     No  nucleus  is  revealed. 

{b)  Water. —  Irrigation  with  water  decolourises  the  corpuscles  and  renders  them  globular. 
The  colouring  matter  is  dissolved  by  the  water  which  passes  into  the  corpuscles  by  endosmosis 
and  renders  them  globular.     The  surrounding  fluid  becomes  yellowish  in  colour. 

{c)  Tannic  Acid. — If  a  drop  of  blood  is  mixed  with  a  two  per  cent,  solution  of  tannic  acid 
(p.  2),  one  or  more  small  refractive  buds  are  seen  to  appear  on  the  margins  of  the  corpuscles. 
These  buds  are  produced  by  the  same  causes  that  produce  similar  buds  on  the  corpuscles  of 
the  newt. 

{d)  Common  Salt. — When  blood  is  irrigated  with  a  two  per  cent,  solution  of  common  salt, 
crenation  of  the  coloured  corpuscles  takes  place — that  is  to  say,  they  become  jagged  at  the 
margins  and  on  the  surface,  so  that  they  resemble  small  thorn-apples.  At  the  same  time,  the 
corpuscles  gradually  become  smaller  and  deepen  in  tint,  but  all  the  corpuscles  are  not  affected 
to   the  same   extent  or  with  the  same  rapidit}-.     Perhaps  this  difference  in  the  action  of  the 


6  PRACTICAL    HISTOLOGY. 

reagent  is  caused  by  some  difference  in  tlie  age  or  physical  properties  of  the  various  cor- 
puscles.    {Indicate  this  in  I'l.  I.,  Fig.  12.) 

(f)  Exposure  to  Air. — If  a  drop  of  blood  be  exposed  to  the  air  before  the  cover  is  applied, 
in  some  cases  crenation  occurs,  but  this  phenomenon  is  not  constant,  nor  when  it  takes 
place  does  it  always  occur  to  the  same  extent. 

None  of  these  preparations  of  human  blood  make  good  permanent  preparations. 

2.  Observe  the  colourless  corpuscles  (H). — In  a  fresh  drop  of  blood  they  are  few  in  number 
—from  five  to  eight  being  usually  in  the  field  at  once.  They  are  best  seen  between  the 
rouleaux  of  coloured  corpuscles,  and  resemble  in  general  characters  the  colourless  corpuscles 
of  the  newt  or  frog.  They  are,  however,  smaller  and  do  not  exhibit  amoeboid  movements 
unless  when  kept  at  the  temperature  of  the  blood  (38°  C.)  on  a  hot  stage  (p.  xxviii).  Some  are 
larger  than  and  others  are  about  the  same  size  as  the  coloured  corpuscles,  and  for  the  most 
part  they  are  all  finely  granular,  though  now  and  again  a  coarsely  granular  one  is  met  with. 
If  the  glass  cover  be  touched  with  the  point  of  a  needle  the  colourless  corpuscles  are  seen 
to  adhere  to  the  glass,  unlike  the  coloured  corpuscles,  which  freely  roll  over  one  another ;  and 
even  when  the  coloured  corpuscles  impinge  on  a  colourless  one  applied  to  the  slide,  they 
seldom  displace  it,  so  firmly  do  these  adhere  to  any  object.  {Indicate  the'  characters  of  the 
colourless  corpuscles  in  PI.  I.,  Fig.  13.) 

EFFECT  OF  REAGENTS. — The  actions  of  Acetic  Acid,  Water  and  Magenta  on  the  colourless 
corpuscles  of  human  blood  are  identical  with  the  effects  of  the  same  reagents  on  the  corre- 
sponding corpuscles  of  the  blood  of  the  newt  or  frog.  {Indicate  the  effect  of  acetic  acid  in 
PI.  I.,  Fig.  14,  and  magenta  in  PI.  I.,  Fig.  15). 

3.  FREE  GRANULES  (H). — Besides  the  two  varieties  of  corpuscles  just  described  (r  and 
2)  a  number  of  very  fine  granules  may  be  observed  in  the  human  blood. 

4.  FIBRIN  (H). — Place  a  cover  on  a  drop  of  blood  on  a  slide  and  allow  it  to  stand  for 
a  quarter  of  an  hour.  At  the  end  of  this  time,  on  careful  examination  a  number  of  delicate 
threads,  which  form  an  open  network  between  the  rouleaux,  may  be  detected.  These  threads 
consist  of  fibrin,  and  are  still  better  seen  if  the  blood  be  irrigated  with  a  drop  of  magenta  or 
iodine  solution.  The  former  colours  the  threads  red,  the  latter  colours  them  yellow.  {Indicate 
the  appearance  of  the  fibrin  threads  in  PI.  I.,  Fig.  9.) 

(C.)     BLOOD-CRYSTALS. 

5.  H.ffiMOGLOBIN. — The  colouring  matter  of  blood,  hemoglobin,  may  be  obtained  in  a 
crystalline  form,  though  it  does  not  occur  in  that  form  inside  the  coloured  corpuscles.  In 
the  corpuscles  it  seems  to  lie  in  the  meshes  of  a  stroma. 

PREPARATION. — Kill  a  guinea-pig  and  take  a  drop  of  blood  from  the  heart  or  else- 
where, mix  it  on  a  slide  with  a  drop  of  distilled  water  and  cover.  The  haemoglobin  diffuses 
from  the  blood-corpuscles  into  the  water,  and,  as  evaporation  takes  place,  needle-shaped 
crystals  form  at  the  margin  of  the  cover-glass.  They  cannot,  however,  be  preserved  for  any 
length  of  time,  so  as  to  make  permanent  preparations.  Or  use  the  blood  of  a  white  rat,  from 
which  hjemoglobin  crystals  form  in  a  few  minutes. 

6.  H.ffiMIN. — Besides  haemoglobin,  which  exists  normally  in  the  coloured  blood-cor- 
puscles, and  from  which  it  may  be   obtained  in  a  crystalline  form,  as  above  described,  certain 


BLOOD.  7 

other  crystals — derivatives  of  haemoglobin — may  also  be  obtained  from  blood.  Of  these  (6), 
Haemin  (Hydrochlorate  of  Hzematin),  or  Teichinaim's  Crystals,  are  of  most  importance,  as  their 
presence  is  considered  one  of  the  surest  as  well  as  one  of  the  most  delicate  tests  for  blood 
in  medico-legal  cases. 

PKEPARATION. — Take  a  drop  of  blood  from  the  finger  and  allow  it  to  dry  on  a  slide,  or 
take  some  powdered  dried  blood  previously  prepared  and  add  to  it  a  trace  of  finely  powdered 
common  salt.  Moisten  the  whole  with  a  drop  of  glacial  acetic  acid,  apply  a  cover,  and  heat 
the  slide  gently  over  the  flame  of  a  spirit  lamp  till  bubbles  of  gas  appear.  Allow  the  prepara- 
tion to  cool,  and  apply  a  cover-glass. 

EXAMINATION.  (H.) — Observe  the  short  reddish-brown  prismatic  crystals  scattered  all 
over  the  field  ;  they  are  best  seen  where  they  are  attached  to  small  masses  of  blood.  These 
are  the  crystals  of  Haemin.  These  make  good  permanent  preparations.  To  preserve  them, 
take  off  the  cover-glass,  and  remove  the  surplus  acid  with  blotting  paper.  Add  a  drop  of 
Tarrant's  solution,  re-apply  the  cover.  For  the  method  of  sealing  up  the  preparation  per- 
manently, see  p.  xlix. 

For  the  method  of  ascertaining  the  number  of  coloured  blood-corpuscles  by  Malassez's 
method,  see  Archives  dc  Physiologic,  1874,  or  the  Practitioner,  July  1S78,  where  Dr.  Gowers 
describes  the  hasmacytometcr.  The  amount  of  haemoglobin  may  be  ascertained  by  means  of 
the  hasmaglobinometer.     See  Clinical  Society's  Transactions,  vol.  xii.,  p.  64. 


PRACTICAL   HISTOLOGY. 


EPITHELIUM. 

Epithelium  consists  of  corpuscles  or  cells  with  or  without  envelopes,  of  various  shapes, 
sizes,  and  consistence,  united  by  an  intercellular  substance  or  cement.  The  corpuscles  are 
always  disposed  on  surfaces,  and  usually  contain  a  nucleus  and  sometimes  a  nucleolus  ;  they 
may  be  colourless,  though  they  sometimes  contain  pigment.  They  may  be  arranged  in  a  single 
layer  or  in  several  layers.  Each  cell  regulates  its  own  nutrition  and  never  contains  a  blood- 
vessel. The  following  is  a  convenient  classification  of  the  varieties  of  epithelium  met  with 
in  the  human  body  : — (i)  Squamous,  (2)  Columnar,  (3)  Ciliated,  (4)  Transitional,  (5)  Secretory. 

I.    SQUAMOUS    EPITHELIUM. 

PREPARATION.— With  the  finger-nail  or  a  blunt  knife  scrape  off  a  little  of  the  epithelium 
lining  the  inside  of  the  cheek  or  covering  the  tongue.  Diffuse  the  scraping  in  water  on  a 
slide,  cover  and  examine. 

EXAMINATION  (H). — In  this  preparation  only  the  superficial  squames  are  seen.  They 
are  irregular  polygonal  cells  when  seen  on  the  flat,  and  are  several  times  the  breadth  of  a 
coloured  blood-corpuscle.  Each  cell  contains  a  relatively  small  nucleus,  and  not  unfrequently 
fine  granules.  Occasionally  several  cells  are  found  adhering  to  each  other  by  their  margins, 
and  sometimes  micrococci  are  seen  attached  to  them.  If  the  edge  of  a  cell  be  directed 
towards  the  observer,  it  will  appear  more  or  less  spindle-shaped  according  to  the  angle  which  it 
forms  with  the  plane  of  the  slide.  {Indicate  these  characters  in  PI.  II.,  Fig.  i.)  In  addition, 
small  round  and  finely  granular  cells,  about  the  size  of  a  colourless  blood-corpuscle  and  con- 
taining one  or  two  nuclei,  may  be  found  here  and  there  in  the  field.  These  are  the  salivary 
corpuscles.  The  included  granules  exhibit  Brownian  movement — a  phenomenon  which  occurs 
when  fine  particles  are  suspended  in  a  watery  fluid,  and  which  is  probably  due  to  differences 
of  temperature  in  the  several  strata  of  the  liquid.  Brownian  movement  is  easily  shown  by 
rubbing  down  a  little  gamboge  in  water. 

EFFECT  OF  REAGENTS.—  Irrigate  with  a  drop  of  magenta  solution  (p.  xliv).  The 
nucleus  stains  of  a  deep  red  colour,  while  the  peri-nuclear  part  is  but  slightly  affected.  The 
alcohol  in  this  solution  precipitates  the  mucin  of  the  saliva  in  the  form  of  fine  red  threads  or 
red  membranous  flakes  (PL  II.,  F"ig.  i). 

Squamous  Epithelium  in  exposed  situations,  as  in  the  superficial  layers  of  the  skin,  becomes 
hardened,  and  forms  what  is  known  as  the  horny  layer  of  the  epidermis. 

PREPARATION. — With  a  knife  shave  off  a  few  of  the  surface  scales  of  the  skin,  diffuse  the 
scraping  in  water,  cover  and  examine. 


EPITHELIUM.  9 

EXAMINATION  (H). — The    squaines    are  flat,  horny,    and    transparent,  and    usually    no" 
nucleus  is  visible. 

EFFECT  OF  REAGENTS.— Irrigate  with  a  five  per  cent,  dilution  of  liquor  potassae;  the  cells 
swell  up  and  become  globular.     Neither  this  nor  the  previous  preparation  need  be  preserved. 

INTRA-CELLULAR    PLEXUS    OF    FIBRILS. 

It  has  recently  been  shown  that  the  nucleus  of  cells  contains  a  delicate  plexus  of  fine 
fibrils — an  intra-niiclear  plexus — which  is  continuous  with  a  similar  plexus  in  the  perinuclear 
protoplasm  — the  intra-cellular  plexus.  It  is  this  plexus,  when  the  ends  of  its  fibres  are  directed 
towards  the  observer,  that  gives  the  cells  the  appearance  usually  described  as  '  granular.' 

PREPARATION.  — Keep  a  newt  in  a  small  quantity  of  water  for  several  days,  but  do  not 
change  the  water.  At  the  end  of  this  time  the  superficial  layer  of  cutaneous  epithelium  will 
slough  off  in  the  form  of  a  fine  film.  Harden  this  film  for  twenty-four  hours  in  a  saturated 
solution  of  picric  acid,  or  in  absolute  alcohol.  Preserve  both  in  alcohol.  When  required  for 
use,  snip  off  a  small  piece  and  stain  it,  if  hardened  in  picric  acid,  with  picrocarmine  for  half 
an  hour,  while  the  piece  hardened  in  alcohol  may  also  be  stained  with  logwood.  Mount  in 
Farrant's  solution. 

EXAMINATION  (H) — Observe  the  cells  united  by  their  margins.  Within  each  polygonal 
area  notice  the  oval  red-stained  nucleus :  study  it  carefully,  and  a  delicate  plexus  of  fibrils — 
the  intra-nuclear  plexus— will  be  seen  (PI.  II.,  Fig.  2). 

Both  in  mucous  membranes  and  in  the  skin,  epithelium  occurs  in  several  Xdiy&x?,,  constituting 
stratified  epithelium.  In  order  to  see  the  deeper  layers,  and  to  study  the  relation  of  one  layer 
to  another,  sections  must  be  made.  A  section  of  the  conjunctival  epithelium  of  the  cornea  is 
convenient  for  this  purpose. 

Vertical  section  of  the  conjunctiva  and  cornea  for  stratified  epithelium. 

PREPARATION.— Remove  the  cornea  from  the  eye  of  a  cat  just  killed  and  place  it  in 
40  c.c.  of  chromic  acid  and  spirit  mixture  (p.  xxxi)  consisting  of  two  parts  of  a  sixth  per  cent, 
solution  of  chromic  acid,  and  one  part  of  methylated  spirit.  Change  the  fluid  at  the  end  of 
the  first,  fourth,  and  seventh  days  ;  on  the  tenth  day  it  will  be  sufficiently  hardened.  For 
preservation  transfer  it  to  methylated  or  rectified  spirit,  after  washing  away  all  the  surplus 
chromic  acid.  When  sections  are  wanted,  put  the  cornea  in  a  large  glass  of  water  for 
twenty-four  hours  to  remove  the  spirit.  Transfer  it  to  a  solution  of  gum  (p.  xxxviii)  for  twenty- 
four  hours  and  then  cut  vertical  sections  in  a  freezing  microtome  (p.  xxxviii).  After  the  sections 
are  made,  they  are  placed  in  a  large  quantity  of  water  for  twenty-four  hours,  to  get  rid  of 
the  gum,  when  they  may  be  put  into  the  preservative  glycerine  fluid  (p.  xl)  till  they  are 
required  for  mounting. 

•EXAMINATION  (H).— Float  a  section  on  to  a  slide  (p.  xxvi),  add  a  drop  of  solution  of 
picrocarmine  (p.  xliii),  and  allow  it  to  remain  on  the  section  for  fifteen  minutes,  or  until  the 
section  is  sufficiently  stained.  Soak  up  the  surplus  staining-fluid  with  blotting-paper,  and  add 
a  drop  of  Farrant's  solution.  Cover  and  examine.  The  fibrous  cornea  is  stained  red,  but 
neglecting  this,  observe  the  layers  of  epithelial  cells,  stained  yellow  and  their  nuclei  red, 
covering  its  anterior  surface.  The  deepest  layer  consists  of  cells  columnar  in  shape  ;  those 
on  the  surface  appear  as  little  more  than  lines  or  very  elongated  spindles,  whilst  between 
these  extremes  there  are  several  layers  of  cells  which  show  intermediate  forms.  The  cells 
are  developed  from  below— from  the  lowest  layer — and  are  gradually  pushed  upwards,  and 
hence  the  change  of  shape.     {Indicate  these  chai-acters  in  PI.  II.,  Fig.  5). 

C 


lo  PRACTICAL    HISTOLOGY. 

2.  COLUMNAR    EPITHELIUM. 

A.     FRESH    COLUMNAR   EPITHELIUM. 

PREPARATION. —  Slit  open  the  small  intestine  of  a  cat  just  killed,  and  wash  away  the 
mucus  covering  its  inner  surface  with  a  stream  of  |  per  cent,  solution  of  common  salt,  scrape 
the  mucous  surface  and  diffuse  the  scraping  in  the  same  strength  of  salt  solution,  and  after 
breaking  it  up  with  needles,  cover  and  irrigate  with  magenta  solution  (p.  xliv). 

EXAMINATION  (H). — A  large  number  of  elongated,  narrow,  or  columnar  cells  are  to  be 
seen.  Each  cell  is  finely  granular  and  contains  a  nucleus  which  is  stained  red.  One  end  of 
the  cell — the  attached  end — is  somewhat  pointed,  while  the  free  end  presents  a  narrow,  clear, 
unstained  hem,  or  border,  marked  with  fine  vertical  lines  or  striae.  This  indicates  that  the 
free  end  of  each  cell  is  covered  by  a  transparent  plate  or  disc.  When  several  cells  adhere  to 
each  other,  and  their  free  ends,  instead  of  their  sides,  are  presented  to  the  observer,  a  fine  mosaic 
is  observed.  The  intercellular  substance  is  small  in  amount,  mapping  out  the  individual  cells 
one  from  another,  while  in  the  centre  of  each  area  a  red  nucleus  is  seen  (PI.  II.,  Fig.  4).  In 
addition,  here  and  there  so-called  'chalice'  or  'goblet'  cells  are  seen  (PL  II.,  Fig.  6).  These 
have  a  cup-shaped  appearance,  are  devoid  of  the  clear  border  and  contain  a  nucleus  sur- 
rounded by  a  small  quantity  of  protoplasm  near  the  lower  pointed  extremity  of  the  cell,  whilst 
the  greater  part  of  the  body  appears  to  be  clear  and  looks  as  if  it  were  empty.  This  is  not 
to  be  preserved  ;  but,  in  order  to  obtain  permanent  preparations,  the  small  intestine  may  be 
treated  in  one  or  other  of  the  following  ways. 

B.     PERMANENT   PREPARATIONS. 

PREPARATION  {a). — Place  a  square  half-inch  of  the  intestine  in  40  to  60  cubic 
centimetres  of  a  one  per  cent,  solution  of  ammonium  bichromate  for  two  days. 

{b)  A  similar  piece  in  dilute  alcohol  (i  rectified  spirit  to  2  water)  for  twenty-four  hours. 

{c)  A  similar  piece  in  a  few  c.c.  (5-10)  of  a  one  per  cent,  solution  of  osmic  acid  (p.  xlvi)  for 
half  an  hour. 

Any  of  these  may  be  used  for  preservation,  but  in  every  case  the  specimen  must  first  be 
steeped  in  water  for  half  an  hour  or  longer,  to  get  rid  of  the  hardening  reagent. 

EXAMINATION  (H). — Scrape  off  a  little  of  the  mucous  surface  of  a  or  b,  place  it  on  a 
slide,  and  break  it  up  with  needles.  Add  a  drop  of  picrocarmine,  and  allow  it  to  stain  for  fifteen 
minutes.     Remove  the  surplus  staining-fluid,  add  a  drop  of  glycerine,  cover  and  examine. 

In  either  case  columnar  and  chalice  cells  similar  to  those  already  described  will  be  found, 
except  that  in  a  each  cell  will  be  yellow  with  a  red  nucleus,  while  in  b  the  red  nucleus  will  be 
seen  to  contain  what  has  been  described  as  a  nucleolus.  If  c  be  similarly  treated,  it  must  be 
left  in  the  picrocarmine  for  twenty-four  hours,  when  the  same  appearances  are  seen  as  in  a, 
with  this  addition — that  if  the  cells  contain  any  fatty  particles,  these  are  blackened.  These 
three  preparations  when  sealed  as  described  (p.  xlix)  are  permanent. 

Columnar  Epithelium  from  the  small  intestine  of  a  newt. — Where  the  cells  are  relatively 
large,  and  show  very  distinctly  the  intra-cellular  and  intra-nuclear  plexus. 

PREPARATION. — Kill  a  newt  and  place  its  small  intestine  in  a  few  c.c.  of  a  five  per  cent. 


EPITHELIUM.  II 

solution  of  ammonium  chromate  for  forty-eight  hours.  After  washing,  transfer  it  to  picro- 
carmine  for  twenty-four  hours.  Open  the  gut,  and  with  a  knife  scrape  off  a  little  of  the 
epithelium  and  diffuse  it  in  a  drop  of  glycerine  on  a  slide,  cover  and  examine. 

EXAMINATION  (H).— Observe  the  tall,  relatively  large,  columnar  cells,  with  a  fine  striated 
disc  on  their  free  ends,  and  a  large  red-stained  nucleus.  A  careful  examination  reveals  the 
plexus  of  fibrils.  Besides  these  cells,  look  for  '  goblet '  or  '  chalice  '  cells,  which  are  somewhat 
cup-shaped,  with  a  nucleus  situated  near  the  narrow,  often  tailed,  end  of  the  cell,  and  surrounded 
by  a  small  quantity  of  protoplasm,  while  the  upper  part  of  the  cell  appears  empty.  These 
cells  secrete  mucin,  so  that  they  may  be  regarded  as  unicellular  glands.  The  mucin  seems  to 
be  derived  from  the  interfibrillar  substance  (PL  II.,  Fig.  7). 


3.     CILIATED    EPITHELIUM. 

A.     LIVING    CILIATED    EPITHELIUM   AND    CILIARY    MOTION. 

PREPARATION. — Kill  a  frog  by  giving  it  a  sharp  blow  on  th^  head,  but  do  not  give 
chloroform,  for  it  paralyses  ciliary  motion.  Scrape  the  mucous  membrane  of  the  roof  of  the 
mouth  or  gullet,  and  diffuse  the  scraping  by  means  of  two  fine  needles  in  J  per  cent,  salt 
solution.     Place  a  hair  in  the  fluid  to  prevent  the  cells  being  crushed,  cover  and  examine. 

EXAMINATION  (H). — Observe  the  groups  of  short  columnar  or  nearly  spherical  cells 
with  very  granular  contents,  and  having  on  one  end  vibratile  cilia  in  active  movement.  This 
movement  causes  currents  in  the  surrounding  fluid,  which  carry  along  with  them  any  suspended 
particles,  such  as  granules  or  blood-corpuscles.  Sometimes  a  detached  cell  may  be  observed 
to  spin  round  and  round  by  the  motion  of  its  own  cilia.  The  cilia  themselves — which  are 
fine  homogeneous  projections — are  best  seen  when  their  movements  become  languid. 

EFFECT  OF  REAGENTS.— Irrigate  with  a  drop  of  magenta  solution,  and  note  that  the 
colouring-matter  does  not  stain  the  cells  until  the  cilia  have  ceased  to  move.  Thus  each  cell 
zoliilc  living  can  regulate  its  own  nutrition.  There  is  a  marked  difference,  therefore,  in  respect 
to  colouring-matters  between  a  living  and  a  dead  cell. 

B.     CILIARY    MOVEMENT. 

For  the  study  of  '  ciliary  nwveinent '  the  cilia  of  the  common  salt-water  mussel  are  best 
adapted.  On  opening  the  mussel  with  a  knife,  the  yellowish-coloured  gills  are  seen,  and  if  a 
small  part  be  snipped  off  and  placed  in  the  salt  water  found  in  the  inside  of  the  shell,  teased 
with  needles,  covered  and  examined  with  a  jow  power — (L) — the  gills  are  seen  as  a  series  of 
bars  fringed  with  cilia,  much  larger  than  those  of  the  frog's  epithelium.  (H).  Observe  the  long, 
tapering,  clear,  and  structureless  cilia  bending  most  at  their  tips.  Seal  up  this  preparation  by 
running  a  ring  of  oil  round  the  margin  of  the  cover-glass,  and  set  it  aside  for  several  hours  till 
the  movement  of  the  cilia  becomes  more  languid  (PI.  II.,  Fig.  10).  The  oil  prevents  the  access 
of  fresh  oxygen,  which  is  necessary  for  the  ciliary  motion  ;  and  as  the  oxygen  in  the  fluid 
becomes  used  up,  the  ciliary  action  becomes  slower.  Place  the  slide  on  a  hot  stage  (p.  x.xviii) 
and  gradually  raise  the  temperature  to  not  higher  than  35°  C.  As  the  temperature  rises,  the 
ciliary  movement  becomes  more  rapid,  but  if  the  temperature  be  too  high,  so  as  to  coagu- 
late the  albumen,  a  permanent  arrest  of  the  movement  results.  An  opposite  effect  is  pro- 
duced by  the  action  of  chloroform  or  ether.     Place  a  fresh  piece  of  the  gill  on  a  cover-glass 


12  PRACTICAL   HISTOLOGY. 

in  a  drop  of  salt-water,  and  invert  it  over  a  gas  chamber  so  that  the  gill  is  directed   towards 
the  chamber  (p.  xxix). 

EXAMINATION  (L). —Ascertain  that  the  cilia  are  active,  then  raise  the  cover-glass,  put  a 
small  drop  of  chloroform  into  the  chamber  by  means  of  a  glass  rod,  and  again  apply  the 
cover-glass.  The  ciliary  movement  becomes  slower  and  slower,  and  finally  ceases.  If,  how- 
ever, the  gill  is  removed  before  the  chloroform  has  acted  too  long,  and  is  freely  exposed  to 
air,  the  cilia  may  resume  their  movements  (Lister).  None  of  these  specimens  can  be  pre- 
served. 

C.     PERMANENT    PREPARATIONS    OF    CILIATED    EPITHELIUM. 

PREPARATION. — Place  a  small  piece  of  the  trachea  of  a  cat,  rabbit,  ox,  or  sheep  in  dilute 
alcohol  (i  to  2  water)  for  forty-eight  hours,  when  the  inter-cellular  substance  of  the  epithelium 
will  be  so  softened  that  the  cells  can  be  easily  isolated.  Then  wash  the  whole  piece  in  water, 
scrape  the  mucous  membrane  with  a  knife,  place  the  scraping  in  a  tube  with  a  few  drops  of  a 
one  per  cent,  solution  of  osmic  acid.  The  isolated  cells  are  thus  '  fixed,'  and  can  be  examined 
either  with  or  without  the  addition  of  various  staining  reagents. 

EXAMINATION  (H).  — Place  some  of  the  cells  in  picrocannine  {ox  at  least  twenty-four 
hours,  transfer  some  to  a  drop  of  glycerine  on  a  slide,  cover  and  examine.  Observe  the 
long,  tapering  character  of  the  cells,  with  one  end  covered  by  a  fringe  of  fine  processes—  the 
cilia — which  are  planted  on  a  clear  disc  which  is  not  stained.  The  cell-protoplasm  is  described 
as  granular,  but  careful  examination  shows  that  it  contains  an  intra-cellular  plexus  of  fibrils 
with  the  fibrils  arranged  chiefly  in  the  long  axis  of  the  cell.  It  is  maintained  by  some 
observers  that  the  cilia  are  actually  prolonged  through  the  clear  band,  so  as  to  become 
continuous  with  this  plexus.  The  oval  nucleus  is  placed  far  down  in  the  cell,  is  bright  red, 
and  shows  a  similar  plexus,  with  a  bright  spot  in  its  centre — the  so-called  nucleolus.  The 
other  end  of  the  cell  is  often  tapered  or  bifurcated  (PL  II.,  Fig.  8).  Instead  of  the  picro- 
carmine  logwood  may  be  used,  with  the  advantage  that  a  few  minutes  suffice  for  the  staining 
process.  Either  of  these  preparations  may  be  sealed  up  and  preserved  permanently.  The 
ends  of  the  cells  may  be  directed  towards  the  observer  (PI.  II.,  Fig.  9). 


4.  TRANSITIONAL   EPITHELIUM. 

PREPARATION. — Take  the  bladder  of  a  sheep,  cat,  rabbit,  or  guinea-pig,  and  prepare  and 
preserve  it  in  the  same  way  as  is  directed  for  the  preservation  of  ciliated  epithelium. 

EXAMINATION  (H). — Observe  the  great  variety  in  the  shape  of  the  cells,  some  of  them 
beino-  more  or  less  cubical,  others  resemble  squames  in  character,  while  others  again  present  a 
number  of  sharp  angular  points  (PI.  II.,  Fig.  1 1). 


5.   SECRETORY   OR   GLANDULAR   EPITHELIUM. 

PREPARATION. — Make  a  cut  into  the  liver  of  a  dog  just  killed  ;  scrape  the  cut  surface 
with  a  knife,  so  as  to  detach  some  of  the  liver-cells.  Place  these  in  a  few  c.c.  of  a  half  per  cent, 
solution  of  osmic  acid  for  an  hour,  then  pour  off"  the  acid  and  substitute  picrocarmine  for 
several  hours. 


X  30  0 


Plate n  Epithelium 


^Arvlem  Bros  litK 


ENDOTHELIUM.  13 

EXAMINATION  (H).  — Many  blood-corpuscles  will  be  seen.  Neglecting  these,  study  a 
liver-cell.  Notice  its  polygonal  shape,  yellowish  granular  protoplasm,  and  its  nucleus  deeply 
stained  red.  No  cell-envelope  is  visible.  Sometimes  there  are  fine  granules  of  pigment  in 
the  protoplasm,  at  other  times  fatty  granules,  especially  if  the  liver  of  a  stall-fed  ox  be 
used.  In  young  animals  (dog)  the  liver-cells  often  contain  two  nuclei.  The  '  granular '  cha- 
racter of  the  nucleus  and  protoplasm  are  due  to  the  existence  of  a  plexus  of  fibrils,  which 
requires  a  higher  magnifying  power  to  make  it  obvious  (PI.  II.,  Fig.  12). 

The  liver-cells  of  the  newt  are  very  large,  and  are  easily  prepared  by  placing  small  pieces 
of  the  liver  in  a  five  per  cent,  solution  of  ammonium  chromate,  and  after  washing,  staining 
them  with  picrocarmine  for  several  hours.  When  teased  out  in  glycerine  they  show  most 
admirably  the  intra-nuclear  plexus  of  fibrils.  The  liver  of  the  newt,  especially  at  the  end  of 
winter,  often  contains  particles  of  pigment  (PI.  II.,  Fig.  12). 


ENDOTHELIUM. 

« 

It  is  convenient  to  apply  this  term  to  a  single  la)er  of  squamous  epithelium  wherever  it 
occurs,  as  the  lining  of  serous  sacs,  blood-  and  lymph-vessels,  &c. 

PEEPARATION  BY  THE  SILVER  PROCESS.— Bleed  a  rabbit,  take  a  portion  of  its  omen- 
tum and  rinse  it  gently  in  distilled  water  to  remove  soluble  chlorides.  Place  it  in  a  quarter 
per  cent,  solution  of  nitrate  of  silver  for  ten  minutes  or  until  it  assumes  a  milky  appearance. 
Wash  it  thoroughly  in  ordinary  water,  and  expose  it  to  the  action  of  diffuse  sunlight,  either  in 
water  or  in  water  and  alcohol,  until  it  assumes  a  brownish  colour.  Mount  a  small  piece 
unstained  in  Farrant's  solution,  and  to  compare  with  it,  stain  a  similar  piece  with  logwood  for 
ten  minutes,  and  after  washing  away  all  the  surplus  logwood,  mount  it  also  in  Farrant's  solu- 
tion. Picrocarmine  does  not  stain  so  well  after  silver,  but  after  a  tissue  has  been  in  alcohol 
for  some  time  it  takes  up  this  pigment  more  easily.  The  omentum  of  the  rabbit  is  chosen 
because  in  it  the  endothelium  occurs  in  a  nearly  continuous  sheet. 

EXAMINATION  (H).— The  tissue  will  be  mapped  out  into  a  series  of  small  polygonal 
areas  by  narrow  black  lines — '  silver  lines.'  These  are  produced  by  a  deposit  of  the  reduced 
silver  in  the  intercellular  cement.  The  flattened  epithelial  cells  adhere  by  their  margins  and 
form  a  complete  epithelium  or  endothelial  investment  for  the  connective  tissue  of  which  the 
omentum  consists.  Focus  carefully  through  the  thickness  of  the  membrane,  and  a  second  set 
of  lines,  whose  outlines  do  not  correspond  with  those  lying  above  them,  will  be  brought  into 
view  ;  these  are  the  endothelial  cells  on  the  other  surface  of  the  membrane  (PI.  II.,  Fig.  3).  It 
is  important  to  practise  this  method  of  focussing  through  the  whole  thickness  of  an  object. 
Usually  no  nucleus  is  observable  within  the  cells,  but  in  the  logwood  preparation  each  cell  will 
be  seen  to  contain  an  oval  nucleus.  Preparations  of  other  serous  membranes  will  be  made 
later  on. 


14  PRACTICAL   HISTOLOGY. 


CARTILAGE. 

In  every  cartilage  there  fall  to  be  considered  {a)  the  matrix,  and  {b)  the  cartilage-ceils  or 
corpuscles.  Cartilages  are  classified  according  to  the  nature  of  the  matrix.  If  the  matrix  be 
clear  and  like  ground-glass  it  is  called  hyaline  cartilage ;  if  fibrous  it  is  a  fibro-cartilage,  of 
which  there  are  two  varieties  :  one,  where  the  matrix  resembles  yellow  elastic  tissue,  is  yellow 
fibro-cartilage,  and  the  other,  in  which  the  fibres  resemble  white  fibrous  tissue,  is  white  fibro- 
cartilage.  Where  the  cells  are  separated  by  a  very  small  quantity  of  matrix,  the  cartilage  is 
said  to  be  cellular,  as  in  the  ear  of  a  mouse  or  in  the  chorda  dorsalis. 


HYALINE    CARTILAGE 

Occurs    in  the   articular  ends  of  bones,  cartilages  of  the  ribs,  part  of  the  sternum,  larynx, 
trachea  and  bronchi,  and  some  nasal  cartilages. 

A.  FRESH  HYALINE  CARTILAGE.— Open  any  joint  of  an  animal  just  killed,  or  take  the 
cartilage  of  a  rib  from  the  same  animal  and  make  a  thin  slice  of  the  cartilage  with  a  razor. 
Place  the  thin  slice  in  a  drop  of  blood-serum  or  one  per  cent,  alum  solution,  cover  and  examine. 
Instead  of  making  a  slice,  a  piece  of  cartilage  sufficiently  thin  may  be  obtained  from  the 
sternum  of  a  newt  or  the  tracheo-laryngeal  cartilages  of  a  frog. 

EXAMINATION  (H). — If  a  thin  slice  has  been  made,  observe  the  hyaline  matrix  dotted 
over  with  corpuscles,  or  cells — the  cartilage-cells.  Study  first  the  cartilage-corpuscles  and 
then  the  matrix.  The  corpuscles  are  oval  or  of  various  shapes,  granular  and  containing  a  large 
nucleus.  The  protoplasm  of  the  cells  fills  the  entire  space  or  lacuna  in  which  it  lies,  but 
— especially  near  the  margin  of  the  section — the  protoplasm  may  have  fallen  out  of  the  lacunae, 
or  shrunk  up  so  that  it  no  longer  completely  fills  the  space  in  which  it  lies.  By  focussing 
the  cells  are  seen  to  lie  in  several  planes.  The  matrix  is  hyaline  and  perfectly  homogeneous 
in  its  appearance  (PI.  III.,  Fig.  i). 

EFFECTS  OF  REAGENTS,  {a)  Water.— Irrigate  the  section  with  distilled  water.  The 
protoplasm  of  the  cell  now  shrinks  rapidly  from  the  cavity  containing  it.  It  becomes 
coarsely  granular  and  thus  often  obscures  the  nucleus. 

{b)  Iodine  Solution. — Make  a  transverse  section  of  a  costal  cartilage  of  a  young  animal. 
Stain  with  iodine  solution  for  three  to  five  minutes,  add  glycerine  and  examine  (H).  This 
stains  the  matrix  yellow,  and  the  corpuscles  of  a  deeper  tint.  Within  the  protoplasm  a 
brownish  coloration  may  result,  which  indicates  the  presence  of  glycogen  ;  this  is  best  seen  in 
young  growing  cartilage. 

B.  METHODS  OF  PREPARING  HYALINE  CARTILAGE  FOR  PERMANENT  PREPARA- 
TIONS.— It  is  necessary  to  use  reagents  to  '  fix '  the  structural  elements  of  the   cartilage,  and 


CARTILAGE. 


'5 


amongst  the  best  of  these  is   a  saturated  watery  solution  of  picric   acid.     Take   costal  and 
articular  cartilage  as  types. 

Place  small  pieces — not  larger  than  half  an  inch — of  the  costal  cartilages,  with  their  peri- 
chondrium, of  a  cat  or  rabbit  or  human  fcetus,  in  a  saturated  watery  solution  of  picric  acid  for 
forty-eight  hours.  Then  remove  them  and  wash  them  thoroughly  in  water  till  no  yellow 
colour  is  given  off,  and  place  them  in  rectified  spirit  till  they  are  required.  Sections  may  be 
made  either  with  the  hand  or  by  means  of  a  freezing  microtome  (p.  xx.xviii).  It  is  well  to 
practise  making  sections  with  a  razor,  and  cartilage,  from  its  consistence,  is  one  of  the  easiest 
tissues  to  begin  with.  A  similar  procedure  is  adopted  for  adult  hitman  costal  cartilage,  and  also 
{ox  articular  cartilage.  If  the  costal  cartilage  contain  any  bony  or  calcareous  matter,  macerate 
it  for  one  or  two  weeks  in  the  acid  till  all  the  calcareous  matter  is  removed.  For  articular  car- 
tilage, if  it  be  desired  to  have  a  section  of  the  subjacent  bone,  the  bone  must  be  softened  by 
means  of  an  acid  which  removes  the  calcareous  matter.  It  may  be  accomplished  thus  : — Place 
the  articular  end  of,  say,  the  femur  or  humerus  of  a  recently  killed  cat  or  rabbit  in  a  large 
quantity  (twenty  to  thirty  times  the  bulk  of  the  tissue)  of  one  of  the  following  solutions: — 

[a)  A  saturated  watery  solution  of  Picric  Acid. — Change  the  fluid  at  the  end  of  a  week  and 
add  fresh  solution  containing  a  few  crj-stals  of  picric  acid.  The  bone  will  be  softened  in  from 
two  to  three  weeks. 

(b)  A  quarter  per  cent,  solution  of  Chromic  Acid  may  be  employed.  Take  care  to  change  the 
fluid  after  three  da)-s.     This  fluid  requires  also  from  two  to  three  weeks  to  decalcify  the  bone. 

{c)  A  mixture  of  Chromic  Acid  and  Nitric  Acid  (p.  xxxiii)  may  be  employed.  Similar  con- 
ditions obtain  as  in  {a)  and  ib). 

{<£)  Or  a  two  per  cent,  solution  of  Hydrochloric  Acid  may  be  used  as  the  decalcifying  agent. 
(«)  and  {c)  are  the  methods  which  yield  the  best  results. 


COSTAL   CARTILAGE. 

Makeanumberof  transverse  sections  of  the  costal  cartilage  of  a  kitten  or  young  rabbit,  and 
also  of  the  decalcified  costal  cartilages  of  an  old  person,  prepared  as  described  above.  These 
sections  may  be  e.xamined  as  they  are  or  after  being  stained. 

A.     FROM   A   YOUNG   ANIMAL. 

{a)  Osmic  Acid. — Place  some  of  the  sections  of  the  kitten's  cartilage  in  a  one  per  cent,  solu- 
tion of  osmic  acid  for  twelve  hours.  Wash  them  thoroughly  in  water,  to  remove  the  osmic 
acid.  Mount  one  in  Farrant's  solution,  and  cover.  These  sections  are  taken,  first,  because 
they  are  small  and  one  sees  better  the  general  characters  of  the  section 

EXAMINATION  (L). — Observe  the  fibrous  perichondrium  and  the  cartilage-matrix  inside 
it,  the  cartilage-corpuscles,  small  and  flattened,  lying  in  several  layers.  Nearer  the  centre  of 
the  section  are  seen,  as  small  specks,  the  irregularly  shaped  corpuscles.  The  corpuscles  are 
stained  of  a  deeper  yellow  than  the  matri.x  by  the  osmic  acid,  hence  its  value.  (H)  Examine 
the  fibrous  perichondrium  with  its  areolar  tissue  and  sections  of  clastic  fibres.  Observe  the 
cartilage-cells  flattened  towards  the  periphery  of  the  cartilage,  and  the  more  rounded  charac- 
ter of  the  cells  placed  nearer  the  centre.  Otherwise  the  cells  and  matrix  have  the  same 
characters  as  are  indicated  in  fresh  hyaline  cartilage. 

(p)  Carmine. — Stain  a  section  with  carmine.     Lay  a  section  on  a  slide  and  on  it  place  a 


1 6  PRACTICAL    HISTOLOGY. 

drop  of  solution  of  strong  carmine  (p.  xliii).  Within  a  few  minutes  it  will  be  stained  of  a 
uniform  deep  red  colour.  Wash  away  the  surplus  carmine,  and  if  the  preparation  is  now  ex- 
amined (L)  it  will  be  of  a  nearly  uniform  red  colour,  the  matrix  scarcely  distinguishable  from 
the  corpuscles  in  the  depth  of  the  tint.  Place  on  the  section  a  large  drop  of  a  five  per  cent, 
solution  of  glacial  acetic  acid  for  several  minutes,  examining  the  section  with  a  low  power  all 
the  time.  The  effect  of  the  acid  is  to  remove  the  surplus  carmine  from  the  matrix  and  leave 
the  corpuscles  deeply  stained  red.  As  soon  as  this  is  accomplished  wash  the  section  thoroughly 
in  water,  and  mount  it  in  Farrant's  solution.  Strong  glacial  acetic  acid  accomplishes  the  same 
result  more  rapidly. 

EXAMINATION  (H).  — The  general  characters  of  the  section  are  the  same  as  («),  only  the 
perichondrium  and  corpuscles  are  red  and  the  matrix  is  colourless  or  only  faintly  stained 
(PL  III.,  Fig.  2). 

B.     FROM   AN    OLD    PERSON. 

Make  a  number  of  transverse  sections  of  the  macerated  tissue  (p.  15).  Let  the  sec- 
tions include  a  piece  which  has  been  calcified. 

{a)  Osmic  Acid. — Place  some  of  the  sections  in  a  one  per  cent,  solution  of  osmic  acid  as 
for  foetal  cartilage. 

EXAMINATION  (L). — Observe  the  perichondrium  and  the  general  arrangement  of  the 
corpuscles,  the  corpuscles  flattened  at  the  periphery,  in  irregular  clumps  a  little  further  in,  and, 
as  one  examines  towards  the  centre,  the  rows  of  cartilage-cells  produced  by  transverse  or 
oblique  cleavage.  The  matrix,  hyaline  at  the  periphery,  but  finely  granular  in  other  parts.  This 
is  best  brought  out  by  slightly  shading  the  mirror  or  using  a  small  aperture  of  the  diaphragm. 

(H).  The  corpuscles — The  protoplasm  is  very  apt  to  shrink  within  the  capsules,  and  in  it 
are  to  be  seen  several  black  spots.  These  are  small  masses  of  oil  which  have  been  blackened 
by  the  osmic  acid,  which  is  an  excellent  reagent  for  detecting  the  existence  of  fatty  particles  in 
any  tissue,  and  is  therefore  of  great  value  In  persons  above  middle  age,  oil-drops  are  fre- 
quently to  be  found  both  in  the  cartilage-cells  of  the  tracheal  rings  and  rib-cartilages.  The 
rows  of  cells  are  obvious. 

Matrix. — Examine  the  granular  part.  Between  the  rows  of  cells  the  matrix  may  be  found 
to  be  distinctly  '  fibrillated  '  ;  the  fibres  lie  mostly  parallel  one  with  another. 

It  is  well  to  mount  a  preparation  of  cartilage,  showing  the  fibrillation  of  the  matrix  in  a 
ten  per  cent,  solution  of  common  salt,  which  prevents  the  swelling  up  of  the  fibrils. 

{U)  Eosin. — Place  other  sections  in  a  dilute  solution  of  eosin  (p.  xlv).  This  substance 
stains  the  section  very  rapidly — a  minute  usually  suffices.  Rinse  them  in  a  one  per  cent,  dilution 
of  acetic  acid  and  mount  in  Farrant's  solution. 

EXAMINATION  (L). — Observe  a  similar  arrangement  of  the  elements.  The  matrix  is 
slightly  red  and  corpuscles  slightly  deeper  in  colour.  The  calcareous  matter  deposited  be- 
tween the  cells  and  the  fibrillated  parts  are  more  deeply  stained  than  the  hyaline  matrix,  and 
a  kind  of  stereoscopic  effect  can  be  given  to  these  calcified  parts  by  using  a  small  aperture  of 
the  diaphragm. 

(H).  Observe  the  highly  refractive  yellow  oil-globules  in  the  protoplasm  of  the  cells,  and 
observe  particularly  the  more  deeply  stained  cell-capsule  or  '  cartilage  capsule '  bounding  the 
lacuna. 

(f)  Purpnrine. — Place  some  sections  in  a  solution  of  purpurine  (p.  xliv)  for  forty-eight  hours. 


Plate  ni  Cartilage 


IfiT-iterr-L  Bros  litK 


CARTILAGE.  17 

This  substance  stains  the  nuclei  of  the  cells  a  light   pink  colour.     It  possesses  no  advantages 
over  the  methods  already  indicated. 

The  advanced  student  may  study  the  action  of  chloride  of  gold  and  nitrate  of  silver  on 
hyaline  cartilage. 

Action  of  Chloride  of  Gold. — Place  in  a  one  per  cent,  solution  of  chloride  of  gold  for  half  an 
hour  the  articular  end  of  the  femur  of  a  frog  newly  killed.  Wash  it  thoroughly  to  remove  the 
surplus  gold,  and  place  it  in  water  acidulated  with  a  few  drops  of  acetic  acid,  and  expose  it  to 
the  light  until  the  gold  is  reduced — usually  for  twenty  hours — or  until  the  cartilage  becomes 
of  a  beautiful  violet  tint.  Make  thin  sections  of  the  cartilage,  and  mount  them  in  Farrant's 
solution. 

EXAMINATION  (H). — Observe  the  cells  arranged  in  several  planes  and  stained  of  a 
deep  violet  colour.  Thej^  usually  fill  the  lacunae  completely.  Often  two  cells  may  be  seen 
together,  in  such  relationship  as  to  suggest  that  they  have  been  produced  by  the  division  or 
'  fission  '  of  a  single  cell.     The  matrix  is  stained  of  a  lighter  colour. 

Action  of  Nitrate  of  Silver. — Silver  the  articular  cartilage  of  the  femur  of  a  freshly  killed  frog, 
as  directed  at  p.  xlv,  and  after  exposure  to  light,  and  when  it  has  become  brown,  make  a  thin 
slice  and  mount  it  in  Farrant's  solution.     Cover. 

EXAMINATION  (H). — Observe  the  matrix  stained  brown,  but  the  cells  are  not  stained, 
they  are  merely  represented  by  oval  or  round  clear  spaces,  which  correspond  exactly  in  shape 
and  distribution  with  the  cartilage-corpuscles.  The  silver  has  not  affected  the  cells,  though  it 
has  darkened  the  intercellular  substance.  The  gold  effect  represents  the  positive,  this  the 
negative  effect.  Staining  with  logwood  reveals  the  presence  of  a  cell  in  each  apparently 
empty  space. 


ARTICULAR   CARTILAGE. 

Make  a  number  of  vertical  sections  of  the  head  and  subjacent  bone  of  the  femur  or 
other  long  bone  of  a  cat  or  other  animal,  decalcified  according  to  the  methods  indicated  (p. 
15),  preferably  by  a  or  c. 

(a)  Osmie  Acid. — Place  sections  in  a  one  per  cent,  solution  of  osmic  acid  for  twenty-four 
hours.  Wash  and  mount  in  glycerine  jelly  (p.  xlviii),  for  Farrant's  solution  renders  the  section 
too  transparent. 

EXAMINATION  (L). — If  the  femur  be  chosen,  observe  the  general  arrangement  of  the 
parts  from  the  articular  surface  downwards.  The  superficial  cells  are  flattened,  and  they  always 
are  parallel  to  the  surface  and  at  right  angles  to  the  axis  of  pressure.  A  little  below  the 
surface  they  are  in  irregular  clumps  of  twos  and  threes,  and  deeper  down  near  the  bone  they 
are  in  rows,  parallel  to  the  long  axis  of  the  bone.  These  rows  are  produced  by  transverse 
cleavage  of  the  cells.  Below  the  cartilage  is  the  bone.  The  cartilage  is  divided  into  two 
zones,  distinctly  mapped  off  from  each  other  by  the  character  of  the  matrix.  Across  the  sec- 
tion, about  two-thirds  or  half-way  below  the  articular  surface  of  the  cartilage,  there  runs  a  line 
slightly  wavy.  All  that  lies  superficial  to  this  has  a  Iiyaline  matrix,  and  what  lies  below  it,  i.e. 
as  far  as  the  bone,  is  more  or  less  granular  and  less  transparent.  In  this  lower  part  the  matrix 
is  calcified.  This  calcified  part  is  distinctly  mapped  off  from  the  subjacent  bone  ;  a  well-marked 
wavy  line  with  very  distinct  undulations — the  one  part  dovetailing  into  the  other,  indicating 

D 


i8  PRACTICAL    HISrOLOGY. 

the  line  of  demarcation.     If  the  section  has  been  made  from  bone  softened  with  chromic  acid 
the  bone  will  have  a  slightly  greenish  tinge. 

(H).  Examine  in  detail  the  characters  of  the  cells,  and  note  especially  the  rows  of  cells  in 
the  deeper  part  of  the  cartilage.  These  rows  are  produced  by  transverse  cleavage  of  the  cells. 
One  part  of  a  row  may  lie  in  the  hyaline  matrix,  while  the  other  half  is  imbedded  in  the 
granular  matrix  (PI.  III.  Fig.  6).  In  some  sections  an  indistinct  fibrillation  is  seen  in  the  lower 
part  of  the  cartilage  between  the  rows  of  cells,  thus  resembling  what  obtains  in  calcifying 
costal  cartilage. 

{b)  Picrocarmine. — Put  a  section  on  a  slide,  and  stain  it  with  picrocarmine  for  twenty 
minutes  to  half  an  hour.  Remove  all  the  picrocarmine,  and  mount  it  in  glycerine  jelly  (p. 
xlviii).  The  glycerine  jelly  must  be  melted  in  a  water  bath,  and  when  fluid  a  drop  is  applied 
to  the  preparation  by  means  of  a  glass  rod,  and  covered  at  once,  before  the  jelly  sets. 


WHITE    FIBRO-CARTILAGE. 

PREPARATION. — Place  a  small  piece  of  an  intervertebral  disc  with  its  adherent  bone,  from 
a  newly  killed  sheep  or  ox  in  twenty  or  thirty  times  its  bulk  of  a  mixture  of  chromic  and 
nitric  acid  (p.  xxxiii).  After  two  to  three  weeks  the  bone  will  be  decalcified.  Wash  the  pieces 
in  water  to  remove  all  acid,  and  preserve  them  in  rectified  spirit,  or  ;— 

Remove  the  skin  from  the  tail  of  a  cat ;  cut  it  into  pieces  an  inch  long,  and  place  these  in 
chromic  and  nitric  acid  fluid. 

Make  with  a  freezing  microtome  vertical  sections  of  the  intervertebral  disc  ;  including 
the  subjacent  bone,  which  will  have  a  greenish  colour.  Make  also  transverse  sections  of  a 
piece  of  the  decalcified  tail  of  a  cat  or  other  animal. 

Place  a  vertical  section  of  an  intervertebral  disc  on  a  slide.  Stain  it  with  strong  carmine 
for  ten  minutes,  or  until  it  assumes  a  deep  red  colour.  Wash,  and  mount  in  Farrant's 
solution. 

EXAMINATION  (L).— Observe  the  relation  of  the  fibrous  part  of  the  disc  to  the  subjacent 
bone. 

(H).  The  matrix  in  the  ox  is  distinctly  fibrous  ;  the  fibres  are  very  fine,  unbranched,  and 
wavy.  The  corpuscles  are  nucleated,  and  enclosed  in  a  distinct  capsule.  They  lie  scattered 
irregularly  or  in  chains  between  the  bundles  of  fibres.  Sometimes  the  cells  are  seen  to 
be  dividing.  Here  and  there  fine  dots,  the  cut  ends  of  the  fibres,  may  be  seen  (PI.  III., 
Fig.  3)- 

Stain  with  carmine  a  transverse  section  of  an  intervertebral  disc  from  the  already 
softened  tail  of  a  cat.  Remove  the  surplus  staining  fluid  with  blotting-paper,  and  mount  in 
Farrant's  solution. 

EXAMINATION  (L). — Observe  the  concentric  arrangement  of  the  parts  of  the  disc  and 
the  different  directions  of  the  fibres  composing  each  layer  of  it. 

(H).  Observe  the  fibrous  character  of  the  matrix  and  the  corpuscles  as  before.  Here 
and  there  are  transverse  sections  of  the  ends  of  the  fibres,  which  appear  like  fine  granules. 


CARTILAGE.  19 


YELLOW   FIBRO-CARTILAGE. 

This  may  be  easily  obtained  from  the  epiglottis,  the  arytenoid  cartilages,  or  the  external  ear. 

PREPARATION. — Place  the  arytenoid  cartilages  and  the  epiglottis  of  a  sheep  or  ox  (if  the 
latter,  cut  it  into  two  pieces)  in  a  saturated  solution  of  picric  acid.  Remove  it  after  forty-eight 
hours  and  make  transverse  sections  of  it  in  the  usual  way.     Preserve  it  in  methylated  spirit. 

Treat  the  external  ear  of  a  cat  or  rabbit  in  the  same  way  and  make  sections.  This 
method  is  preferable  to  simple  hardening  in  alcohol. 

Stain  a  transverse  section  of  an  epiglottis  hardened  as  above  with  picrocarmine.  A  few 
minutes  suffice  to  stain  the  parts.  Remove  the  surplus  staining  fluid  with  blotting  paper, 
and  mount  in  Farrant's  solution. 

EXAMINATION  (L). — This  specimen  contains  a  section  of  the  mucous  membrane  covering 
both  surfaces  of  the  epiglottis.  This  may  be  neglected  at  present.  The  cartilage  itself  is 
stained  bright  yellow  (picric  acid)  with  a  perichondrium  of  connective  tissue  which  is  of  a  deep 
red  colour.  Lying  in  the  bright  yellow  fibrous  matrix  are  a  number  of  red  spots,  which 
are  the  nuclei  of  the  cartilage-corpuscles. 

(H).  Notice  the  wrt//'/.!:- -yellow,  distinctly  fibrous,  and  the  fibres  like  those  of  elastic  tissue, 
and  branched.  If  the  line  of  .section  cuts  these  fibres  transversely  they  appear  as  fine  yellow 
granules.  Trace  these  fibres  into  the  red  connective-tissue  perichondrium,  where  they  become 
yellow  elastic  fibres.  This  continuity  can  be  easily  made  out  in  a  thin  section.  Study  the 
corpuscles.  Towards  the  periphery  of  the  cartilage  they  are  flattened  ;  but  near  the  centre 
they  are  rounded,  have  a  distinct  capsule,  and  the  nucleus  is  distinctly  stained  red.  This 
forms  an  extremely  beautiful  preparation.  These  preparations  require  a  day  or  two  for  the 
colours  to  become  differentiated. 

A  section  may  be  double-stained  with  picrocarmine  and  logwood,  but  this  method  pos- 
sesses no  advantages  over  the  above. 

Stain  a  section  of  the  ear  of  a  cat  with  picrocarmine  as  above.    Mount  it  in  Farrant's  solution. 

EXAMINATION  (L  and  H). — The  yellow  mass  of  cartilage  stands  out  beautifully  between 
the  red  connective  tissue  (Pi.  III.,  Figs.  4  and  $).  Sections  of  muscles,  skin,  and  sebaceous 
glands  may  also  be  seen.  The  same  general  characters  are  seen  with  a  high  power  as  are  indi- 
cated above. 

TRANSITION    FROM    HYALINE    TO    YELLOW    FIBRO-C.A.RTILAGE. 

Make  longitudinal  sections  of  an  arytenoid  cartilage  of  an  ox,  and  stain  it  with  picro- 
carmine. The  upper  half  shows  the  .structure  of  yellow  fibro-cartilage,  while  the  lower  half  is 
chiefly  hyaline,  with  a  few  elastic  fibres  extending  into  the  matrix.  This  shows  the  mode  of 
development  of  yellow  fibro-cartilage  from  hyaline  cartilage. 

CHANNELS    IN    THE    MATRIX    OF    HYALINE    CARTILAGE. 

These  are  easily  seen  in  a  section  of  a  part  of  the  cephalic  cartilage  of  Loligo,  which  has 
been  hardened  in  picric  acid  and  stained  with  picrocarmine. 


20  PRACTICAL   HISTOLOGY. 


CONNECTIl^E    TISSUE. 

Connective  tissue  occurs  in  a  variety  of  forms,  as  areolar  tissue,  or  membranes,  or  forming 
fascia;  and  tendons.  In  whatever  form  it  occurs  we  have  to  study  two  classes  of  structural  ele- 
ments, v\z.—fibi-cs  and  corpuscles.  The  fibres  are  of  two  kinds,  [a)  the  zvliite  or  gelatigejwns,  so 
called  because  they  yield  gelatine  on  boiling  ;  (/;)  the  yellow  or  elastic,  which  yield  elastin.  The 
corpuscles  are  of  three  kinds  :  {a)  the  fixed  connective-tissue  corpuscles,  which  always  bear  a 
definite  relation  to  the  white  fibres;  (b)  auiceboid  ox  zcawc/tv/;/^  ctV/y,  which  are  identical  with 
colourless  blood-corpuscles  or  leucocytes.  They  wander  freely  in  the  lymph-spaces  by  virtue 
of  their  contractility ;  {c)  the  so-called  plasma  cells,  of  which  the  exact  relation  is  not  made  out. 


ELASTIC   TISSUE. 

A.     IN    THE    FRESH    STATE. 

PREPARATION. — Tear  off,  in  the  axis  of  the  fibres,  with  a  forceps,  a  very  fine  piece  of  the 
fresh  ligamentum  nuchae  of  an  ox.  Tease  it  out  in  a  drop  of  salt  solution  or  water,  with 
two  needles,  and  cover.  It  is  convenient  to  use  a  low  power  to  ascertain  when  the  piece  is 
sufficiently  teased. 

EXAMINATION  (H).—  Notice  the  homogeneous  transparent  fibres,  about  the  breadth  of  a 
human  coloured  blood-corpuscle.  They  have  a  yellowish  tinge,  and  their  margins  are  well- 
defined.     They  branch  and  anastomose  and  may  curl  up  at  the  ends  (PI.  IV.,  Fig.  i). 

EFFECT  OF  REAGENTS. — Irrigate  the  preparation  with  a  one  per  cent,  dilution  of  acetic 
acid.  The  fibres  are  not  affected  thereby.  To  preserve  this  preparation  substitute  Farrant's 
solution  for  the  acid  watery  fluid. 

B.     PERMANENT   PREPARATIONS. 

Excellent  permanent  preparations  are  obtained  by  the  following  method.  Place  small 
pieces — about  half-inch  cubes — in  many  times  their  bulk  of  chromic  and  nitric  acid  fluid 
(p.  xxxiii)  for  a  week.  After  that  soak  the  pieces  in  water  for  twenty-four  hours,  to  remove  all 
traces  of  the  acid,  and  preserve  them  for  use  in  methylated  spirit.  Make  longitudinal  and 
transverse  sections  in  the  usual  way. 

Stain  a  longitudinal  and  a  transverse  section  in  picrocarmine  for  ten  minutes. 

EXAMINATION  (H)  of  the  longitudinal  section  reveals  the  elastic  fibres  stained  of  a  deep 
yellow,  with  their  characteristic  branches  and  anastomoses.  Between  them  a  small  amount 
of  ordinary  connective  tissue  stained  red  is  seen. 

The  transverse  section  shows  the  cut  ends  of  the  fibres,  as  solid  yellow  polygonal  bodies  in 


CONNECTIVE   TISSUE.  2r 

groups  of  twos  and  threes  ;  when  two  fibres  adhere  their  opposed  surfaces  are  moulded  to 
each  other  (PI.  IV.,  Fig.  2).  Surrounding  each  group  of  fibres  is  a  very  small  quantity  of 
connective  tissue  stained  red.  Only  a  very  small  quantity  of  white  fibrous  tissue  exists  in 
the  ligamentum  nucha;.  If  desired  similar  sections  may  be  tinged  with  a  solution  of  magenta 
(p.  xliv),  which  stains  the  elastic  fibres  bright  red,  and  is  thus  a  good  reagent  for  detecting 
their  presence. 

Elastic  tissue  also  forms  part  of  ordinary  areolar  tissue.  It  occurs  in  the  form  of  a  net- 
work of  fine  clastic  fibres  in  the  skin,  where  it  will  be  examined,  but  its  characters  may  be 
well  studied  in  the  mesocolon  of  a  rabbit. 

PREPARATION. —Remove  the  mesocolon  from  a  newly  killed  rabbit.  Pin  it  to  a  piece 
of  flat  cork  to  keep  it  stretched,  and  place  it,  cork  uppermost,  for  three  days  in  a  mixture  of 
chromic  and  nitric  acid  (p.  xxxili).  After  washing  away  all  surplus  acid,  preserve  it  in  methy- 
lated spirit.  Snip  off  a  small  piece,  float  it  in  water  to  remove  the  spirit,  and  stain  it  for  ten 
minutes  with  picrocarmine  ;  mount  it  in  Farrant's  solution. 

EXAMINATION  (H). — Observe  the  network  of  fine  elastic  fibres  one-sixth  the  breadth  of 
a  coloured  blood-corpuscle.  The  fibres  are  imbedded  in  white  fibrous  tissue,  which  is  stained 
red. 

If  desired  another  piece  may  be  stained  with  solution  of  magenta  (p.  xliv)  which  stains  the 
elastic  network  red.  Elastic  tissue  will  be  further  considered  with  arteries,  and  in  the  skin, 
where  the  method  of  preparing  it  by  artificial  digestion  is  described  (Stirling)  (p.  92J. 


WHITE    FIBROUS   TISSUE. 

IN    THE    FRESH    STATE. 

PREPARATION. — Kill  a  rabbit  or  a  rat,  and  with  scissors  snip  off  a  small  piece  of  the 
delicate  connective  tissue  that  lies  under  the  skin,  and  which  connects  it  to  the  fascia;.  The 
piece  of  tissue  at  once  rolls  into  the  form  of  a  ball  ;  but  by  placing  it  on  a  dry  slide  it  can 
easily  be  spread  out  with  needles  into  a  fine  membrane,  its  original  form  when  in  situ. 
During  the  process  do  not  allow  the  tissue  to  dry,  which  may  be  prevented  by  breathing  on  it 
from  time  to  time.     Add  a  drop  of  normal  salt  solution  ;  cover  and  examine. 

EXAMINATION  (H). — Observe  the  wavy  fibres  of  white  fibrous  tissue  running  in  every 
direction.  Their  outlines  are  indistinct,  but  here  and  there  a  longitudinal  striation  in  their 
course  is  observable,  indicating  that  each  fibre  is  composed  of  fibrils.  In  addition  to  these,  a 
few  fibres  which  refract  the  light  more  strongly  may  be  detected  ;  these  are  elastic  fibres  ;  but 
little  else  is  obserAable  in  this  preparation,  especially  if  taken  from  an  adult  animal  (PI.  IV., 
Fig.  3)- 

EFFECT  OF  REAGENTS,  (a)  Magenta  Solution.— Irrigate  the  preparation  with  solution 
of  magenta  (p.  xliv),  and  the  existence  of  the  corpuscular  element  will  be  revealed.  The  nuclei 
of  the  connective-tissue  corpuscles  are  stained  of  a  red  colour  by  the  dye.  There  are  two  kinds 
of  corpuscles,  one  larger  than  the  other.  The  larger,  when  seen  on  the  flat,  are  like  large 
nucleated  squames,  though  they  are  sometimes  branched,  and  when  seen  on  edge  they  are 
fusiform.  These  are  the  '  fixed  '  connective-tissue  corpuscles,  and  the  smaller  ones,  about  the 
size  of  the  colourless  blood-corpuscles,  with  two  or  three  small  nuclei,  are  the  wandering  cells, 
or  leucocytes,  and  are  identical  with  lymph-  or  colourless  blood-corpuscles.     The  elastic  fibres 


2  2  PRACTICAL    HISTOLOGY. 

— which  arc  recognised  b}-  their  branching  and  forming  a  network — are  stained  red,  while  the 
vhite  fibres  are  but  shghtly  acted  on  by  the  magenta.  This  preparation  need  not  be  pre- 
served. The  observer  is  not  to  suppose  that  the  corpuscular  elements  do  not  exist  before  the 
addition  of  a  reagent  ;  they  do  exist,  but,  owing  to  their  refractive  index  being  so  near  that  of 
the  fluid  in  which  they  are  examined,  they  are  not  visible  until  some  staining  reagent  is  added. 
It  is  further  to  be  remembered  that  the  relation  of  the  fixed  corpuscles  to  the  white  fibres  is 
changed  in  a  teased  preparation.  Occasionally  large  oval  granular  and  nucleated  cells  are 
found,  especially  along  the  course  of  the  vessels  ;  they  are  the  so-called //^ri-ww  cells. 

(d)  Dilute  Acetic  Acid.  —  Make  a  similar  preparation,  and  irrigate  it  with  a  one  per  cent, 
dilution  of  acetic  acid. 

EXAMINATION  (H). — Note  that,  as  the  acetic  acid  displaces  the  salt  solution,  it  acts 
upon  the  white  fibres  and  causes  them  to  swell  up,  to  become  transparent,  so  that  they  appear 
quite  homogeneous  and  jelly  like,  though  they  are  not  dissolved.  Some  of  the  fibres  may, 
however,  present  annular  constrictions  in  their  course.  This  clearing  up  of  the  white  fibres 
brings  the  other  elements  in  the  tissue  more  clearly  into  view — to  wit,  the  elastic  fibres  and 
the  corpuscles.  The  elastic  fibres  are  highly  refractive,  and  their  branches  unite  to  form  a  net- 
work, and  if  such  a  fibre  be  broken  across  it  curls  up  at  its  ends,  though  otherwise  they  pursue 
a  straight  course.  Numerous  fusiform  or  oval  nuclei  are  revealed  ;  they  are  the  nuclei  of  the 
fixed  connective-tissue  corpuscles,  whilst  the  small  bi-  or  tri-partite  nuclei  belong  to  the  lymph- 
corpuscles.  The  acetic  acid  may  be  displaced  by  magenta  solution,  when  the  elastic  fibres 
and  the  nuclei  are  stained  of  a  blight  red  colour  (PI.  IV.,  Fig.  4).     Do  not  preserve  this. 


AREOLAR    TISSUE. 

PERMANENT    PREPARATIONS    OF   AREOLAR   TISSUE. 

PREPARATION  (a). — With  a  small  subcutaneous  syringe  (fig.  27)  forcibly  inject  a  quantity 
of  a  half  per  cent,  solution  of  osmic  acid  under  the  skin  of  the  groin  of  a  young  dog,  rat,  or 

rabbit   just    killed,    until    an    artificial 
Fig.  27.  oedema  or  bulla  is  formed.     After  half 

an  hour,  snip  open  the  bulla,  taking 
care  that  no  hairs  get  into  it.  With 
scissors  snip  off  a  small  piece  of  the 
distended  connective  tissue  and  place 
it  on  a  slide.  Spread  it  out  as  a  mem- 
brane with  needles,  and  add  picrocar- 
mine.      It   will    require    several   hours 

Subcutaneous  syringe  for  interstitial  injections,  and  the  cannula.     The    to    stain,  and  to  avoid    desiccation    add 
hollow  needle  ought  to  be  made  of  gold,  to  prevent  acids  from  acting  ,,     ,  ^      1  ■  , 

on  it.    Natural  size.  a  small  drop    of  glycerme  and  cover. 

When  it  is  sufficiently  stained,  remove 
the  picrocarmine,  and  substitute  glycerine  containing  a  trace  of  formic  acid. 

(d)  A  similar  preparation  may  be  stained  with   logwood — which   acts  in   a   few  minutes  ; 
remove  the  logwood  solution  and  mount  in  glycerine. 

(c)  Instead  of  injecting  osmic  acid,  the  subcutaneous  injection  of  a  saturated  watery  solution 
of  picric  acid  does  very  well.     Stain  the  section  with  picrocarmine,  as  directed  under  (<5). 


CONNECTIVE   TISSUE.  23 

EXAMINATION  (H). — The  corpuscles  are  now  clearly  brought  into  view  ;  their  nuclei  are 
stained  and  their  wing-shaped  expansions  are  distinct, — in  fact,  they  appear  as  flat,  nucleated 
plates,  sometimes  witlj  divided  expansions.  Elastic  fibres  are  yellow,  and  white  fibres  are 
slightly  reddish. 

Annular  Constrictions  on  the  White  Fibres. — Take  a  small  piece  of  the  delicate  pia  mater 
lying  within  the  circle  of  Willis  from  the  brain  of  a  sheep  just  killed,  or  slit  up  the  dura 
mater  of  the  spinal  cord  of  an  ox,  and  remove  some  of  the  fine  fibres  that  stretch  across  the 
sub-arachnoid  space.  Spread  out  the  tissue  on  a  slide,  and  irrigate  it  with  a  one  per  cent, 
dilution  of  acetic  acid.  The  fibres  swell  up,  but  here  and  there  they  exhibit  numerous  annular 
constrictions.  These  constrictions  are  the  indications  of  the  existence  of  a  sheath  on  the 
fibres,  which  is  not  affected  by  acetic  acid.  In  some  cases  a  fibre  may  be  found  to  wind  in  a 
spiral  manner  round  the  white  fibre.  These  preparations  may  be  stained  with  picrocarmine 
or  magenta,  but  they  do  not  keep  well. 

The  fibres  of  white  fibrous  tissue  are  made  up  of  fibrils,  held  together  by  a  cement. — The 
fibrils  (juflTHT  inch  ''i  diameter)  are  sometimes  revealed  by  teasing,  but  it  is  better  to  use  some 
chemical  reagent  to  dissolve  the  cement. 

PREPARATION. — Place  a  narrow  strip  of  the  tendo  Achillis  or  other  tendon  of  a  rabbit 
or  other  animal  in  a  saturated  solution  of  picric  acid  (p.  xxxii)  for  twenty-four  hours  After 
that  time  wash  away  the  surplus  colouring  matter,  tear  off  a  small  shred,  and  tease  it  in  water. 
Cover. 

EXAMINATION. — The  wavy,  excessively  delicate,  non-branched,  yellow-coloured  fibrils 
(due  to  picric  acid)  are  now  apparent.  Each  white  fibre  is  made  up  of  a  great  number  of 
these  fibrils,  held  together  by  a  cement.  Usually  no  cells  are  seen,  but  irrigation  with  a  one 
per  cent,  dilution  of  acetic  acid  readily  brings  the  long  oval  nuclei  of  the  tendon-cells  into  view. 

OTHER  METHODS. — Maceration  of  a  piece  of  tendon  in  a  saturated  watery  solution  of 
baryta  for  four  or  five  hours,  and  subsequent  steeping  in  water  for  twenty-four  hours — or 
m.aceration  in  a  large  quantity  of  a  ten  per  cent,  solution  of  common  salt  for  ten  days, 
effect  a  sim.ilar  result,  but  the  picric  acid  method  is  the  most  satisfactory  and  convenient. 


TENDON. 

It  is  advisable  to  use  a  tendon  from  a  young  animal,  for  in  such  there  is  a  much  larger 
number  of  corpuscles  than  in  the  tendon  of  an  adult. 

PREPARATION. —  Place  pieces  of  the  tendo  Achillis  of  a  calf— about  an  inch  long  and  a 
quarter  of  an  inch  thick — in  many  times  their  bulk  of  Muller's  fluid  (p.  xx.xii)  for  fourteen  days. 
Then  remove  them,  and  after  soaking  them  in  water  to  remove  the  chromium  salts,  preserve 
them  in  rectified  spirit  till_  sections  require  to  be  made  in  the  usual  way  b}'  freezing.  Make 
both  transverse  and  longitudinal  sections. 

TRANSVERSE    SECTION. 

(l)  Place  a  section  on  a  slide  and  cover  it  with  a  large  drop  of  logwood  solution  ; 
within  a  few  minutes  it  will  be  sufficiently  stained.     Wash  away  all  the  surplus  logwood  and 

mount  the  section  in  Farrant's  solution. 


24  PRACTICAL    HISTOLOGY. 

EXAMINATION  (L).— Observe  the  sheath  of  the  tendon,  composed  of  connective-tissue 
fibres  disposed  circularly.  It  sends  processes  into  the  substance  of  the  tendon,  subdividing  it 
into  polygonal  areas.  These  areas  are  made  up  of  the  cut  ends  of  the  longitudinally  disposed 
fibres  which  constitute  the  tendon.  Each  area  has  scattered  in  it  a  number  of  small  specks 
deeply  stained  by  the  logwood,  united  by  fine  lines.  These  are  the  spaces — the  inter-fascicular 
lymph-canals— between  the  bundles  of  fibres,  and  in  each  space  a  corpuscle   may  be  detected 

(PI.  IV.,  Fig.  6). 

(H).  The  great  mass  of  the  tendon  is  but  slightly  stained,  but  notice  the  sections  of  the 
corpuscles,  which  are  deeply  tinted.  The  corpuscles  give  off  processes  which  may  be  traced 
a  certain  distance  between  the  fibre-bundles.  Amongst  the  cut  ends  of  the  fibres  may  be 
seen  small,  light,  refractive  points  ;  these  are  the  ends  of  elastic  fibres— few  in  number— 
which  exist  in  tendon  (PI  IV.,  Fig.  7). 

The  branched  intcrfasciadar  spaces  in  the  transverse  section  are  best  seen  in  a  transverse 
section  of  a  dried  tendon  examined  in  water.  The  spaces  appear  branched  and  black  and 
larger  than  natural,  from  the  shrinking  of  the  bundles  of  fibres  bounding  them.  They  are 
black  because  they  contain  air.  The  addition  of  glycerine  renders  all  the  parts  too  trans- 
parent, hence  such  a  preparation  is  not  permanent. 

Sometimes  the  logwood  stains  the  section  of  an  almost  uniform  dark  violet  tint,  and  it 
becomes  impossible  to  distinguish  the  corpuscles  from  the  rest  of  the  section.  Under 
these  circumstances,  irrigate  the  over-stained  section  with  a  five  per  cent,  dilution  of  glacial 
acetic  acid,  examining  the  section  all  the  time  with  a  low  power.  The  colouring  matter  will 
be  dissolved  out  of  the  fibres  and  remain  in  the  corpuscles,  and  if  the  acid  is  thoroughly 
washed  out  of  the  preparation,  this  may  be  mounted  in  Farrant's  solution  and  preserved.  It  is 
well  to  place  such  a  .section  for  a  few  minutes  in  a  five  per  cent,  solution  of  bicarbonate  of 
soda  to  remove  all  the  acid.  This  process  of  overstaining  a  section  with  either  logwood  or 
carmine,  and  getting  rid  of  the  excess  by  means  of  acetic  acid,  is  frequently  of  value. 


LONGITUDINAL    SECTION. 

(2)  Place  a  section  on  a  slide  and  stain  it  with  logwood,  as  directed  for  a  transverse  sec- 
tion. 

EXAMINATION  (H). — Observe  the  fibres  arranged  longitudinally,  many  of  them  slightly 
wavy,  their  outlines  indistinct  and  always  unbranched.  Between  the  fibres,  which  are  but 
slightly  stained  or  not  at  all,  rows  of  elongated  fusiform  corpuscles  deeply  stained  are 
observed.  Study  these  corpuscles.  They  are  nucleated  and  the  nucleus  is  surrounded  by  a 
granular  mass  of  protoplasm  which  forms  part  of  the  body  of  the  corpuscle.  They  are  fixed 
connective-tissue  corpuscles  or  '  tendon  cells,'  and  are  most  numerous  in  young  tendons 
(PI.  IV.,  Fig.  5). 

(3)  Tease  out  with  needles  a  small  piece  of  a  section  treated  as  (2).  The  fibres  and  cor- 
puscles are  easily  isolated.  Notice  the  nucleated  granular  corpuscles  in  many  cases  adhering 
to  and  partly  encircling  a  fibre.  These  corpuscles  when  seen  on  edge  are  fusiform,  but  when 
detached  and  seen  on  the  flat  they  are  more  or  less  flattened,  with  their  edges  sometimes  turned 
in.     These  cells  partly  surround  and  form  an  imperfect  sheath  for  the  fibres. 

Ranvier  has  shown  that  the  relations  of  the  cells  to  the  fibres  of  a  tendon  can  be  more 
successfully  made  out  in  the  fine  tendons  found  in  the  tail  of  a  young  rat  or  mouse. 


X  300 


Plate 'iv:  Connective 


% 


r 


Mrtx.iem  Sros  lith 


CONNECTIVE    TISSUE. 


TENDONS    IN    THE   TAIL   OF   A    RAT    OR   MOUSE. 

PREPAEATION. — Kill  a  rat — preferably  a  young  one — seize  the  tip  of  the  tail,  and  for- 
cibly pull  it.  This  ruptures  the  caudal  vertebrae,  and  a  leash  of  fine  silvery-looking  tendon.s 
is  pulled  out.  Place  several  of  these  tendons  in  the  filtered  juice  of  a  lemon.  Leave  them 
there  for  three  minutes.  They  become  clear  and  transparent  and  swell  up  considerably. 
Remove  them,  wash  them  thoroughly  in  water,  and  place  them  in  several  times  their  bulk  of 
a  one  per  cent,  watery  solution  of  chloride  of  gold  (p.  xlv),  and  allow  them  to  remain  there 
for  fifteen  or  twenty  minutes.  They  become  of  a  deep  yellow  colour,  contract  somewhat, 
and  are  rendered  more  rigid.  Remove  them  from  the  gold  solution,  and  wash  away  the  sur- 
plus gold  in  water.  Place  them  in  an  ounce  of  a  twenty-five  per  cent,  watery  solution  of 
formic  acid.  The  bottle  containing  them  must  be  kept  in  a  dark  place  for  twenty-four  hours. 
At  the  end  of  this  time,  the  tendons  are  removed  from  the  formic  acid  and  are  thoroughly 
washed  in  water.  They  have  now  a  decided  purple  colour  throughout.  The  formic  acid  re- 
duces the  gold  on  which  it  is  deposited  in  the  tissues,  and  to  a  greater  extent  in  the  protoplasm 
of  the  cells  than  in  the  fibres.  There  are  other  ways  of  employing  the  '  gold  method,'  but  we 
have  found  this  modification  of  Ranvier's  very  successful.  [Each  student  cannot  do  the  entire 
process  ;  but  he  is  shown  how  to  do  it.]  Take  a  small  piece  of  one  of  the  above  tendons, 
and  tease  it  with  needles,  in  a  drop  of  glycerine,  and  apply  a  cover-glass,  or  what  does  very 
well,  gently  press  on  the  cover-glass  until  the  tendon  is  flattened. 

EXAMINATION  (H). — Observe  the  rows  of  flattened  quadrilateral  cells  lying  between  the 
longitudinally  disposed  fibres.  The  central  part  of  each  cell  which  contains  the  nucleus  is 
of  a  dark  purple  colour  and  finely  granular.  Thin  slightly  coloured  wings  may  be  seen  ex- 
tending outwards  from  this.  The  position  of  the  nucleus  is  often  left  unstained  and  is  usually 
placed  at  one  end  of  the  cell,  and  the  nuclei  of  adjacent  cells  in  a  row  are  usually  so  disposed 
that  the  two  nuclei  lie  close  together.  One  or  more  dark  lines  may  be  seen  running  along  the 
centre  of  the  cells  (Boll's  stripe).  These  are  ridges  projecting  in  a  direction  vertical  to  the 
plane  of  the  rectangular  plate  of  the  cells  (PL  IV.,  Fig.  8).  It  is  easy,  by  teasing  out  the 
preparation  carefully,  to  isolate  a  fibre  with  some  of  the  cells  still  adhering  to  and  partly 
embracing  it,  so  as  to  indicate  the  relation  of  the  cells  to  the  fibres.  They  can  be  seen  to 
send  their  thin  lamellar  or  wing-like  expansions  around  the  fibre,  so  as  partly  to  embrace 
it  (Pi.  IV.,  Fig.  9).  These  cells  partly  occupy  the  branched  interfascicular  spaces  seen  in  a 
transverse  section  of  tendon  (p.  24). 


DOUBLE-STAINING    OF   TENDONS. 

Gold  Chloride  and  an  aniline  dye.— Place  an  inch  of  the  tail  of  a  rat  in  gold  chloride  for  an 
hour,  and  then  treat  it  as  above.  After  the  gold  is  reduced  place  the  tail  in  chromic  and  nitric 
acid  fluid,  to  remove  the  lime-salts  (p.  xxxiii).  Make  transverse  sections,  and  stain  them  with 
iodine  green  and  aniline  blue,  and  mount  them  in  dammar  (p.  1).  Beautiful  sections  of  tendons 
stained  with  gold  are  obtained,  while  part  of  the  bone  or  interstitial  cartilage  becomes  green. 
These  form  most  instructive  preparations. 

CELL-SPACES    IN    TENDON. 

The  above-described  cells  lie  in  spaces,  which  may  be  demonstrated  by  a  solution    of 

E 


26  PRACTICAL   HISTOLOGY. 

nitrate  of  silver — just  as  in  cartilage  (p.  17)  ;    the  silver  blackens  the  intercellular  parts  and 
leaves  the  spaces  occupied  by  the  cells  clear. 

PREPARATION. — Take  several  of  the  freshly  exposed  tendons  of  the  tail  of  a  rat.  Wash 
them  with  a  camel-hair  pencil  dipped  in  distilled  water.  Do  this  several  times  to  remove 
the  epithelium  which  covers  their  surface.  Place  the  tendons  in  a  half  per  cent,  solution  of 
nitrate  of  silver  for  five  minutes.  Then  wash  them  in  water  and  mount  a  small  piece  of  a 
tendon  so  treated  in  glycerine  ;  and  expose  it  to  the  light  until  it  assumes  a  brownish  colour. 

EXAMINATION  (H). — Observe  the  irregular  spaces  seen  in  a  dark  brown  matrix.  These 
spaces  communicate  one  with  another  by  fine  clear  lines.  The  brown  stained  part  indicates 
the  existence  of  an  intercellular  substance ;  in  the  clear  spaces  lie  the  cells  with  their 
processes. 

EPITHELIAL   COVERING   OF   TENDONS. 

Each  of  the  fine  tendons  is  enveloped  by  a  layer  of  squamous  epithelium,  which  is  easily 
demonstrated  by  staining  the  tendons — witlwut  p)xvious  pencilling — with  nitrate  of  silver  as 
directed  above.  Mount  a  tendon  in  glycerine,  and  expose  it  to  light  until  it  becomes  brownish 
in  colour. 

EXAMINATION  (H). — The  outlines  of  the  polygonal  epithelium  are  distinctly  seen.  These 
indicate  the  existence  of  an  epithelial  investment  of  the  tendon  superficial  to  the  fibres  and 
cells  already  described. 


MEMBRANOUS   CONNECTIVE    TISSUE. 

THE    OMENTUM. 

A  convenient  form  of  this  tissue  is  found  in  the  omentum,  which  differs  in  its  characters 
in  various  animals,  and  also  in  young  and  in  adult  animals.  In  an  adiclt  cat  the  omentum 
consists  of  bundles  of  connective  tissue  of  various  sizes,  cemented  to  each  other  in  the  same 
plane,  so  as  to  form  a  network.  In  the  larger  trabeculas  blood-vessels  and  fat-cells  are  found. 
Each  bundle  of  fibres  is  completely  invested  by  a  layer  of  squamous  epithelium  or  endothelium, 
and  the  nuclei  of  these  cells  can  readily  be  seen,  on  the  surface  or  edges  of  the  fibres  when  they 
are  stained.  The  trabeculae  consist  of  fibrils  which  can  be  seen  in  the  fresh  tissue.  In  the 
substance  of  the  trabeculae  and  amongst  the  fibrils  a  few  fusiform  corpuscles — the  connective- 
tissue  corpuscles — may  be  seen.  They  are  relatively  few  in  number,  and  so  are  the  elastic 
fibres. 

In  a  young  grozving  rabbit  the  omentum  is  much  less  fenestrated,  and  forms  a  more 
continuous  sheet  of  connective  tissue,  covered  on  each  surface  with  a  layer  of  endothelial 
squames.     As  the  animal  becomes  older  the  membrane  becomes  more  fenestrated. 

PREPARATION.  Silver  Nitrate. — Remove  the  omentum  from  an  adult  cat  and  rabbit, 
and  also  from  a  young  rabbit  just  killed.  Wash  each  in  distilled  water  to  remove  the  soluble 
chlorides,  and  then  place  them  in  a  quarter  per  cent,  solution  of  nitrate  of  silver  for  five  minutes, 
and,  after  washing  them  thoroughly,  expose  them  in  alcohol  to  the  light  till  they  become  brown. 
Snip  out  a  small  piece  of  each  with  scissors,  and  float  them  from  water  on  to  slides  in  the  usual 
way  (p.  xxvi).  Mount  in  Farrant's  solution  and  cover,  or  similar  preparations  may  be  stained 
with  logwood,  which  stains  the  nuclei. 


CONNECTIVE   TISSUE.  27 

EXAMINATION  (L  and  H). — Observe  the  nearly  continuous  layer  of  endothelium  in  the 
case  of  the  young  rabbit  (PI.  V.,  Fig.  i)  on  each  surface  of  the  omentum  (p.  26),  the  more 
fenestrated  character  in  the  adult  rabbit,  and  the  almost  net-like  condition  in  the  cat  (PI.  V., 
Fig.  2).  In  all  cases  the  silver  lines,  indicating  the  existence  of  the  endothelium  covering 
the  trabeculae,  are  seen.  Focus  down  through  one  of  the  larger  trabeculae,  inside  which  con- 
nective-tissue corpuscles  may  be  found.  Do  not  confound  the  large  oval  nuclei  of  the  squames 
which  are  always  on  the  surface  with  the  nuclei  of  the  connective-tissue  corpuscles  proper. 
Omit  at  present  the  study  of  the  blood-vessels  and  the  fat  cells  which  frequently  accompany 
them. 

The  omentum  of  a  young  rabbit  is  particularly  valuable  for  studying  the  development  of 
fat-cells  and  blood-vessels. 


ADIPOSE    TISSUE. 

This  consists  of  small  vesicles  or  cells  filled  with  fatty  substances.  The  cells  are  imbedded 
in  a  small  quantity  of  areolar  tissue  containing  blood-vessels.  Each  cell  has  a  distinct  en- 
velope, and  contains  a  globule  of  clear,  sometimes  slightly  yellow-coloured  fat,  which  nearly 
fills  the  cell,  so  that  the  nucleus,  with  a  small  amount  of  protoplasm  surrounding  it,  is  pushed 
to  one  side  and  compressed  against  the  cell-wall.  Study  fully  formed  cells  and  fat-cells 
developing. 

PREPARATION. — Snip  off  a  small  piece  of  fat  and  tease  it  in  a  drop  of  salt  solution  on  a 
slide.  Place  a  piece  of  paper  between  the  slide  and  cover-glass,  to  prevent  the  latter  pressing 
on  and  rupturing  the  envelopes  of  the  cells. 

EXAMINATION  (L  and  H).— Observe  the  relatively  large  size  (3^  -  ■^)  of  the  clear,  highly 
refractive  vesicles,  sometimes  compressed  against  each  other.  Usually  no  nucleus  is  visible. 
If  the  cells  have  been  ruptured,  globules  of  fat  of  various  sizes  will  be  found  floating  about. 
Irrigate  with  magenta  solution  (p.  xliv),  which  stains  the  nuclei  of  the  cells.  They  are  seen 
as  small  oval  red  spots  compressed  against  the  cell-envelope.  Substitute  glycerine  for  the 
magenta,  and  mount  for  preservation.     {Indicate  this  in  PI.  V.,  Fig.  4.) 

Effect  of  Osmic  Acid.— A  similar  preparation,  without  tlie  magenta,  may  be  irrigated  with 
a  one  per  cent,  solution  of  osmic  acid,  which  blackens  fatty  matter,  and  hence  is  a  most  valuable 
reagent  for  its  detection. 

Examine  the  omentum  of  the  cat  (p.  26)  and  the  preparations  of  areolar  tissue  (p.  22) 
for  fat-cells. 

Crystals  of  Margarine. — Delicate,  needle-shaped  crystals,  springing  from  a  point  within  the 
fat-cells,  often  occur.  They  are  post-mortem  appearances — for  the  contents  of  fat-cells  are 
fluid  during  life — and  may  readily  be  produced  by  steeping  a  morsel  of  fat  for  twenty-four 
hours  in  glycerine,  or  long  steeping  in  alcohol,  or  after  partial  digestion  (p.  92). 

The  Envelope  of  Fat  Cells  is  easilj'  shown  by  steeping  a  morsel  of  fat  in  ether,  which  dis- 
solves out  the  fat  and  leaves  the  empty  shrivelled-up  envelopes.  {Indicate  tins  in  PI.  V.^ 
Fig.  5.) 

DEVELOPMENT  OF  FAT-CELLS. 

PREPARATION. — Place  a  small  piece  of  the  skin  of  a  foetus  in  rectified  spirit  for  three 
days.  At  the  end  of  that  time,  make  vertical  sections  in  the  ordinary  way  by  freezing. 
After  the  gum  has  been  removed  from  them  by  steeping  in  water,  place  some  of  them  in  a 

E  2 


28  PRACTICAL   HISTOLOGY. 

small  quantity  of  a  one  per  cent,  solution  of  osmic  acid  for  five  hours.     Wash  them  thoroughl)^ 
in  water,  and  mount  one  in  Farrant's  solution.     Cover. 

EXAMINATION  (L). — Note  the  skin  and  the  subcutaneous  tissue,  and,  scattered  through- 
out the  latter,  various  sized  groups  of  black  spots.  These  black  spots  are  of  various  sizes, 
and  are  the  fat-cells,  which  have  been  blackened  by  the  osmic  acid.  Select  a  group  and 
examine.     {Indicate  this  general  arrangenient  in  PL  V.,  Fig.  3.) 

(H).  Notice  the  different  sizes  of  the  various  vesicles  composing  each  group.  Some  of 
the  vesicles  appear  to  be  nearly  filled  with  a  blackened  globule  of  fat,  whilst  others  contain 
only  one  or  two  small  black  granules  of  fat,  the  rest  of  the  cell  being  occupied  with  a  finely 
granular  protoplasm.  These  cells  are  connective-tissue  corpuscles  in  process  of  being  trans- 
formed into  fat-cells  through  the  degeneration  of  their  protoplasm  into  fat.  Developing  fat- 
cells  may  be  well  studied  in  the  connective  tissue  of  the  infra-orbital  region  of  a  young  rabbit. 

PIGMENT    IN    CONNECTIVE-TISSUE    CORPUSCLES. 

Connective-tissue  corpuscles  containing  pigment  are  seen  in  the  skin  of  the  frog  or  in  its 
mesentery,  and,  in  fact,  in  nearly  all  its  tissues.  Preparations  will  be  obtained  from  the 
choroid  coat  of  the  eyeball  of  any  animal  (p.  107). 


MUCOUS   TISSUE. 

Mucous  or  embryonal  tissue  has  a  close  relation  to  adipose  tissue,  as  in  the  embryo  it 
forms  the  subcutaneous  tissue  which  ultimately  becomes  panniculus  adiposus  in  the  adult. 
It  is  essentially  an  embryonal  tissue,  as  in  addition  to  the  site  mentioned  above,  it  occurs  only 
in  the  umbilical  cord  (Wharton's  jelly),  and  in  the  vitreous  humour  of  the  eye.  Acetic  acid 
precipitates  the  mucin  in  it  in  the  form  of  fine  threads.  It  consists  of  fusiform  and  branching 
cells,  whose  processes  communicate  in  all  directions  with  each  other,  so  as  to  form  a  mesh- 
work,  whose  meshes  are  filled  with  a  clear  transparent  mucin-yielding  ground  substance.  In 
it  also  are  flat  endothelial  cells  similar  to  those  found  in  connective  tissue.  It  soon  changes 
its  character  as  the  embryo  grows  older  ;  white  fibrous  tissue  appears  in  the  matrix,  so  that 
it  comes  partially  to  resemble  connective  tissue,  and  then  to  pass  into  adipose  tissue.  In  the 
umbilical  cord  this  last  change  does  not  take  place  :  it  remains  in  the  foetus  at  full  time  as  a 
fibrous  tissue  containing  a  mucin-yielding  fluid  in  its  meshes. 

PREPARATION  {a).  Subcutaneous  Mucous  Tissue. — Inject  into  the  axilla  or  groin  of  as 
young  an  embryo  as  it  is  possible  to  get  a  small  quantity  of  a  quarter  per  cent,  solution  of 
osmic  acid,  so  as  to  form  a  bulla.  Snip  out  a  small  part  of  the  cedematous  tissue,  stain  it 
with  logwood  or  picrocarmine,  and  mount  in  glycerine. 

EXAMINATION  (L  and  H).— Observe  the  fusiform  and  also  the  branched  cells  anastomo- 
sing with  each  other  with  their  nuclei  stained.     Use  a  very  small  aperture  of  the  diaphragm. 

{U)  Umbilical  Cord. — Harden  part  of  the  umbilical  cord  of  a  four  months'  foetus  in  Midler's 
fluid  for  a  week,  and  make  transverse  sections.  Stain  a  section  with  logwood,  and  mount 
it  in  Farrant's  solution.  Another  may  be  stained  with  methyl-aniline  (p.  xliv)  and  mounted 
in  a  saturated  solution  of  acetate  of  potash. 

EXAMINATION  (L  and  H).— Observe  the  three  umbilical  vessels  (p.  124)  surrounded  by 


5  no 


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CONNECTIVE    TISSUE.  29 

mucous  tissue  as  above  described.  The  cells  are  larger  and  the  meshwork  wider  in  the  part 
farthest  from  the  vessels.  The  intercellular  substance,  even  at  this  age,  contains  connec- 
tive-tissue fibres,  and  their  number  increases  in  older  specimens  ;  hence  the  necessity  for  having 
as  young  a  foetus  as  possible  to  see  the  uncomplicated  characters  of  this  tissue.  (^Indicate  these 
corpuscles  in  PI.  V.,  Fig.  6.) 

If  a  similar  preparation  be  teased  with  needles   and  stained  with   methyl-aniline  before 
it  is  mounted,  it  is  easy  to  isolate  flattened  endothelial  plates  with  wing-like  expansions. 


ADENOID   TISSUE. 

This  tissue  forms  the  basis  of  the  lymphatic  glands  and  the  spleen,  and  occurs  widely 
distributed  as  simple  lymphatic  glands  or  in  diffuse  patches  in  many  organs  of  the  body.  It 
was  formerly  believed  to  consist  of  branched  corpuscles,  whose  processes  anastomosed  with 
the  processes  of  adjoining  cells,  so  as  to  form  a  network,  which  was  filled  with  lymph-cor- 
puscles. We  now  know,  however,  that  it  is  built  on  the  same  type  as  connective  tissue.  It 
consists  of  a  ground  substance,  dense  reticulum  or  network — adenoid  reticulmn — made  up  of 
excessively  fine  homogeneous  fibrils  and  membranes,  which  form  a  honeycomb-like  open 
network.  Transparent,  flattened  connective  tissue  or  endothelial  cells  with  oval  flattened 
nuclei  are  applied  here  and  there  to  these  fibres,  and  partially  embrace  or  enclose  them  just 
as  in  connective  tissue  The  large  nuclei  which  usually  occur  at  the  nodes  of  the  network, 
and  which  were  formerly  regarded  as  the  nuclei  of  the  branched  cells,  are  really  the  nuclei  of 
the  flattened  cells  applied  to  the  fibres.  This  meshwork  is  filled  with  lynipJi-corpuscles. 
These  are  small  cells  exactly  like  colourless  blood-corpuscles,  with  a  small  round  nucleus, 
which  is  thus  easily  distinguished  from  the  large  oval  nucleus  of  the  endothelial  plate.  They 
are  not  all,  however,  of  the  same  size  At  first  sight  only  these  lymph-corpuscles  are  seen, 
and  so  densely  packed  are  they  that  one  requires  to  shake  or  pencil  them  out  of  the  mesh- 
work before  the  adenoid  reticulum  can  be  brought  into  view. 

PREPARATION  {a). — Place  a  small  piece  of  a  fresh  lymphatic  gland  of  an  ox,  or  an  entire 
gland  from  the  abdominal  cavity  of  a  cat,  dog,  or  rabbit,  for  a  week  in  Muller's  fluid.  Make 
transverse  sections.  Stain  a  section  deeply  with  logwood,  and  then  place  it  in  a  test-tube 
with  water  and  shake  it  briskly  for  five  minutes  or  so.  This  detaches  the  lymph-corpuscles, 
and  the  previous  staining  enables  one  to  see  the  section.  Float  it  on  to  a  slide,  and  mount  in 
Farrant's  solution. 

{U)  Nitrate  of  Silver  Injection  and  Logwood. — Into  a  fresh  mesenteric  or  lymphatic  gland, 
thrust  the  nozzle  of  a  hypodermic  syringe  filled  with  a  quarter  per  cent,  silver  nitrate  solution, 
and  push  in  the  silver  solution  into  the  gland  so  as  to  form  an  artificial  cedema.  The  gland 
must  be  tinged  with  the  silver  solution.  Place  it  for  twenty-four  hours  in  alcohol  and  then 
make  sections  by  freezing.  At  first  the  sections  will  be  colourless  ;  but  on  exposure  to  light, 
wherever  the  silver  has  penetrated,  they  will  be  stained  brown.  Stain  a  section  in  logwood, 
and  mount  it  in  Farrant's  solution  or  dammar.     See  also  lymphatic  gland  (p.  "JJ^ 

EXAMINATION  (L). —  Indistinct  indications  of  the  adenoid  reticulum  here  and  there 
crowded  with  lymph-corpuscles  stained  blue  will  be  seen.  Select  a  spot  where  {^^n  corpuscles 
are  to  be  seen  and  examine  it  with  (H).  Study  the  excessively  delicate  adenoid  reticulum, 
with  here  and  there  a  large  oval  nucleus  lying  on  it,  especially  near  a  node.     These  are  the 


30  PRACTICAL   HISTOLOGY. 

nuclei  of  the  endothelial  plates.  Observe  the  small  lymph-corpuscles  in  the  meshwork.  Not 
unfrequcntly  in  the  mesenteric  glands  of  the  ox  certain  of  the  lymph-cells  will  be  found  to 
contain  a  brownish-coloured  pigment,  and  I  have  found  multi-nucleated  cells  closely  resembling 
those  found  in  the  spleen.  The  source  of  the  pigment  in  the  one  case,  and  the  origin  and  function 
of  these  multi-nucleated  cells,  have  still  to  be  investigated.  In  the  silver  preparation  certain 
parts  acted  on  by  the  silver  are  brown,  while  those  unacted  on  are  bluish  from  the  logwood 
stain.  The  silver  opens  up  the  network  somewhat,  and  thickens  the  fibrils  of  which  it  is 
composed,  so  that  it  is  more  easily  seen.  It  forms  an  instructive  preparation.  {Indicate  the 
characters  of  adenoid  tissue  in  PI.  V.,  Fig.  7.) 


BONE. 

PREPARATION.  Methods  of  softening  bone,  {a)  Chromic  and  nitric  acid. — Dissect  out  a 
long  bone — the  femur  or  humerus  of  a  cat  or  rabbit — remove  the  muscles  attached  to  it,  but 
retain  the  periosteum.  With  a  saw  cut  the  bone  into  pieces  three-quarters  of  an  inch  in 
length,  and  place  them  in  a  bottle  with  300  cubic  centimetres  of  one-sixth  per  cent,  solution 
of  chromic  acid  (p.  xxxi).  After  three  days  substitute  a  quarter  per  cent,  solution  of  chromic 
acid,  and  after  three  days  more  a  half  per  cent,  for  another  three  days.  Then  transfer  the 
pieces  of  bone  to  a  large  quantity  of  a  mixture  of  chromic  and  nitric  acid  (p.  xxxiii).  The 
object  is  to  remove  the  calcareous  matter  and  leave  the  structure  of  the  bone  intact.  The 
chromic  acid  alone  '  fixes '  the  elements  and  partially  decalcifies  the  bone,  but  to  employ 
chromic  acid  solution  alone  requires  a  long  time  to  complete  the  decalcification,  hence  the  use 
of  chromic  and  nitric  acid  mixture,  which  completes  the  process  more  rapidl}',  with  no  risk  of 
the  more  delicate  structures  in  the  bone  being  injured  by  the  action  of  the  nitric  acid.  It 
expedites  the  process  to  change  the  fluid  frequently.  At  the  end  of  three  weeks  the  bones 
still  retain  their  form,  though  flexible  and  soft,  and  can  now  be  cut  with  a  knife.  By  thrusting 
a  needle  into  the  bone  one  can  determine  when  all  the  calcareous  matter  has  been  removed,  there 
being  an  absence  of  grittiness.  The  bone  will  now  have  a  greenish  colour,  from  the  deposition 
of  a  chromic  sesquioxide.  Place  the  bone  in  a  large  quantity  of  water,  frequently  changed  to 
get  rid  of  the  acids.  Either  make  sections  (transverse  and  longitudinal)  at  once  in  the  usual 
way,  or  place  the  bones  in  alcohol  till  they  are  required. 

(b)  Picric  acid  method. — Place  similar  pieces  of  bone  in  a  saturated  solution  of  picric  acid 
and  add  a  few  crystals  of  the  acid  to  keep  the  solution  saturated,  as  part  of  the  acid  is  used  up 
to  combine  with  the  lime  in  the  bone.  This  process  takes  much  longer  time  than  the  above, 
but  it  answers  admirably  for  foetal  bones  or  for  very  small  bones.  After  decalcification  wash 
the  bones  for  a  long  time  in  water,  to  get  rid  of  the  intensely  yellow  colouring  matter.  Place 
them  in  spirit  till  they  are  required.  Make  transverse  sections  of  the  shaft  of  a  bone,  including 
the  periosteum,  decalcified  in  either  of  the  above  ways.  Place  some  of  the  sections  in  a 
one  per  cent,  solution  of  osmic  acid  for  twelve  hours.  Wash  them  thoroughly  and  preserve 
them  in  weak  spirit. 

{c)  A  10  per  cent,  solution  of  sodic  chloride  and  hydrochloric  acid. — This  is  for  showing  the 
fibrillar  structure  of  the  bone-matrix  (v.  Ebner  and  De  B.  Birch).  This  method  is  fully  de- 
scribed at  p.  xxxiii.    The  sections  must  be  preserved  in  a  ten  per  cent,  solution  of  sodic  chloride. 

{d)  Artificial  digestion  with  trypsin. — See  p.  xxxiv. 


OSSEOUS   TISSUE.  31 


T.  S.    SHAFT   OF   A   DENSE   BONE    DECALCIFIED. 

EXAMINATION  (L).  {a)  Osmic  acid  section. — Float  the  section  on  to  a  slide  from  weak 
spirit,  remove  the  surplus  spirit,  add  a  drop  of  melted  glycerine  jelly  (p.  xlviii),  and  cover. 
(Glycerine  and  Farrant's  solution  render  the  tissues  too  transparent.)  Observe  the  periosteum 
surrounding  the  ring  of  bone.  If  the  section  be  made  through  the  centre  of  the  shaft  there  will 
be  little  cancellated  tissue — only  a  ring  of  dense  bone  surrounding  the  medullary  cavity.  In 
the  bone  observe  the  Haversian  canals,  some  divided  transversely,  others  obliquely,  and  some 
are  opened  into  longitudinally  ;  arranged  with  reference  to  these  are  the  bone-corpuscles  lying  in 
their  lacunae,  which  appear  like  little  dark  specks.  Some  lacuna;  are  arranged  with  reference 
to  the  periphery  of  the  bone.  The  lamella;  of  bone  may  be  faintly  seen.  Those  arranged  round 
the  Haversian  canals  are  the  Haversian  lamellae,  and  these  with  the  lacunie  and  their  cana- 
liculi  and  Haversian  canal  make  up  a  Haversian  system,  the  peripheric  lamellae  arranged 
with  reference  to  the  periphery  of  the  bone,  and  lastly  some  segments  of  larger  circles  which 
lie  jammed  up  between  the  Haversian  systems,  though  not  belonging  to  them — the  interme- 
diary lamellae,  which  are  segments  of  greater  circles  than  the  Haversian  lamellae.  This  in- 
dicates that  at  one  time  they  were  situated  under  the  periosteum,  but  in  process  of  develop- 
ment they  have  come  to  lie  amongst  the  Haversian  systems.  They  never  contain  a  Haversian 
canal.     {Indicate  these  general  characters  in  PI.  VI.,  Fig.  I.) 

(H).  Observe  the  outer  and  inner  layers  of  the  periosteum  and  perhaps  a  layer  of  somewhat 
flattened  cells  lying  directly  on  the  bone  between  it  and  the  periosteum — the  osteoblasts. 
Observe  the  bone-matrix  with  the  faintly  indicated  lamellae,  and  the  bone-corpuscles — irregular, 
somewhat  shrunken,  nucleated  masses  of  protoplasm,  each  lying  in  a  space  or  /aciina,  with  faint 
indications  of  the  canaliculi.  The  Haversian  canals,  opened  into  longitudinally,  may  be  seen 
to  contain  osteoblasts  and  a  blood-vessel.     {Indicate  these  structures  in  PL  VI.,  Fig.  2.) 

{b)  Stain,  a  T.  S.  of  the  shaft  with  picrocarmine  for  ten  minutes,  and  mount  it  in  Farrant's 
solution  or  glycerine  jelly.  Cover.  (L).  The  periosteum  is  now  well  defined,  its  outer  layer 
being  stained  red,  from  its  consisting  chiefly  of  white  fibrous  tissue,  whilst  the  deeper  layer  is 
somewhat  yellower  in  colour.     The  matrix  of  the  bone  is  yellow  and  the  bone-corpuscles  red. 

(H).  The  outer  layer  of  the  periosteum  is  now  seen  to  consist  of  bundles  of  fibrous  tissue 
crossing  each  other,  mixed  with  a  few  elastic  fibres  ;  the  deeper  yellow-coloured  layer  consists 
of  a  large  proportion  of  elastic  fibres,  mostly  arranged  longitudinally,  and  so  their  cut  ends 
appear  as  dots,  and  lying  on  the  bone  is  a  layer  of  flattened  nucleated  cells  stained  yellow — the 
osteoblasts. 

L.  S.    OF   A    DENSE   BONE    DECALCIFIED. 

EXAMINATION  (L). — Observe  the  Haversian  canals  cut  into  and  arranged  for  the  most  part 
longitudinally,  with  here  and  there  transverse  or  oblique  canals  connecting  them  ;  the  rows  of 
lacunae— five  or  more  between  two  Haversian  canals  ;  the  lacuna;  arc  flattened,  with  their  long 
axis  in  the  axis  of  the  bone,  and  from  each  surface  they  give  off  canaliculi.  If  the  periosteum 
has  been  preserved  the  osteoblasts  may  be  traced  into  a  Haversian  canal  opening  on  the 
surface  of  the  bone.     {Iidicate  the  Haversian  canals  and  lacjinw  in  PI.  VI.,  Fig.  3.) 

(H).  Observe  the  lacunae  with  their  included  bone-corpuscles  ;  in  the  Haversian  canals 
the  remains  of  blood-vessels  and  osteoblasts. 


PRACTICAL    HISTOLOGY. 


BONE    LAMELL/E. 

PREPARATION. — {a)  With  forceps  pull  off  a  thin  layer  of  the  peripheric  lamellae  from  a 
bone  softened  in  dilute  hydrochloric  acid  (p.  xxxiii)  and  denuded  of  its  periosteum.  Examine 
in  water.  If  it  is  to  be  preserved,  it  must  be  placed  for  twenty-four  hours  in  one-half  per 
cent,  osmic  acid,  and  mounted  in  Farrant's  solution. 

{b)  Digestion  of  a  piece  of  bone  with  artificial  pancreatic  juice  (p.  xxxiv),  previously  softened 
with  dilute  chromic  acid,  enables  one  to  isolate  the  fibres  of  which  the  lamella;  arc  composed. 
(Birch.) 

The  sections  must  be  preserved  in  a  ten  per  cent,  solution  of  sodic  chloride. 

EXAMINATION  (H). — The  thin  sheet  of  tissue  consists  of  fibres  resembling  white  fibrous 
tissue,  which  appear  to  cross  each  other  in  two  directions,  and  here  and  there  a  hole  may  be 
seen,  through  which  a  Sharpey's  fibre  had  passed.  Projecting  from  the  surface  may  be  seen 
finger-like  processes  ;  these  are  Sharpey's  fibres,  which  have  been  pulled  out  of  their  sockets. 

SHARPEY'S   FIBRES. 

PREPARATION. — Decalcify  a  piece  of  a  human  parietal  bone  in  dilute  hydrochloric  acid. 
Make  vertical  sections,  and  place  one  on  a  slide.  With  needles  tear  off  the  peripheric  lamellae, 
using  a  dissecting  microscope  (p.  xxxv).  Some  of  the  fibres  will  thus  be  torn  from  their 
sockets,  and  will  project  as  fine  processes  from  the  lamellae. 

EXAMINATION  (L  and  H). — Observe  the  fine  elongated  nail-like  process,  and  perhaps 
the  socket  from  which  they  were  removed. 

In  the  foregoing  preparation  of  a  lamella  they  are  seen  directed  towards  the  observer. 
No  Sharpey's  fibres,  such  as  are  described  above,  are  found  within  the  Haversian  systems. 
They  are  found  in  the  peripheric  and  intermediary  lamellae. 

ELASTIC   FIBRES    IN    BONE. 

In  addition  to  these  fibres  of  white  fibrous  tissue,  elastic  fibres  also  pass  into  the  bone 
from  the  periosteum.  These  are  best  seen  in  transverse  sections  of  a  decalcified  shaft  of  a 
long  bone,  which  has  been  slightly  stained  with  glycerine  to  which  a  trace  of  magenta  solution 
has  been  added  (magenta-glycerine).  The  magenta  stains  these  fibres  of  a  deep  red,  while 
the  white  fibres  are  unaffected  by  the  dye.  Further,  the  elastic  fibres  branch,  which  the 
white  never  do.  The  magenta-glycerine  must  be  allowed  to  act  slowly,  and  when  the  section 
is  sufficiently  stained,  remove  it,  and  substitute  for  it  Farrant's  solution,  and  cover. 


FCETAL   BONE. 

This  ought  to  be  softened,  preferably  with  picric  acid  solution  (p.  xxxii).  Select  a  long 
bone  of  a  human  foetus  or  the  bones  of  a  newly  born  kitten.  They  require  a  shorter  time  for 
decalcification  than  adult  bones.  Make  transverse  sections  and  preserve  them  in  preservative 
fluid  (p.  xl)  until  required.  Stain  a  section  with  picrocarmine,  and  mount  it  in  Farrant's  solu- 
tion and  cover. 


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OSSEOUS   TISSUE.  ^t, 


T.    S.    OF    DENSE    FCETAL    BONE    (DECALCIFIED). 

EXAMINATION  (L).— The  bone-matrix  is  yellowish  and  the  corpuscles  in  the  lacunae  red. 
Observe  the  ring  of  bone,  not  so  dense  as  in  the  adult ;  the  Haversian  canals  are  relatively 
larger,  and  contain  a  layer  of  osteoblasts,  which  line  them,  and  also  a  blood-vessel.  The 
outer  la)-er  of  the  periosteum  is  red  and  the  deeper  layer  is  yellowish,  and  between  it  and  the 
bone  a  layer  of  nucleated,  somewhat  cubical  cells — the  osteoblasts — which  may  be  traced  into 
Haversian  canals  opening  on  the  surface  of  the  bone.  (H).  Observe  the  osteoblasts  lying 
on  the  bone,  and  here  and  there  they  may  be  traced  into  Haversian  canals  which  run 
horizontally  and  communicate  with  the  surface  of  the  bone.  The  bone-corpuscles  are  relatively 
larger_  than  in  adult  bone.  After  a  few  days,  they  become  quite  red  and  are  better  defined. 
The  canaliculi  may  also  be  traced.  Under  the  periosteum  giant-cells  may  be  found  here  and 
there.     {Indicate  the  periosteum,  the  osteoblasts,  and  bone-corpuscles  in  PI.  VI.,  Fig.  6.) 


DENSE    DRY    BONE. 

PREPARATION. — Take  a  human  radius  which  has  been  completely  freed  from  grease  ;  with 
a  fret-saw  cut  a  very  thin  transverse  section.  Grind  the  section  on  a  hone  moistened  with  water. 
Examine  with  a  low  power  to  determine  when  the  section  has  been  ground  thin  enough. 
Rinse  it  in  water  to  get  rid  of  the  debris,  and  allow  it  to  dry.  Take  Canada  balsam,  which  is 
quite  hard  in  the  cold,  and  place  a  small  piece  of  it  on  a  slide,  gently  warmed  over  the  flame  of  a 
spirit-lamp  till  the  balsam  melts.  Allow  the  balsam  to  cool,  and  before  it  has  become  quite 
hard,  place  on  it  the  section  of  bone,  and  cover.  This  method  prevents  the  balsam  from 
getting  into  the  lacunce  and  canaliculi.  and  so  rendering  the  bone  too  transparent.  Make  a 
corresponding  longitudinal  section  of  the  shaft  of  a  long  bone. 

T.   S.   OF    DENSE    DRY   BONE. 

EXAMINATION  (L).—  Observe  the  Haversian  systems,  each  consisting  of  a  Haversian 
canal  surrounded  by  lamelL-e  disposed  concentrically  with  relation  to  it— Haversian  lamellae  ; 
amongst  these  are  seen  the  lacuna;  which  resemble  black  dots  with  fine  branches— the 
canaliculi— proceeding  from  them.  Observe  also  the  circumferential  or  peripheric,  and  the 
intersystemic  or  intermediary  lamellse. 

(H).  Observe  the  shape— flattened  ovals— and  arrangement  of  the  lacuna;  ;  they  appear 
black,  being  filled  with  air.  From  them  proceed  fine,  branching  canals,  which  appear  as  fine 
black  lines,  running  across  the  lamellae  and  uniting  the  lacunae  by  a  network  of  very  fine 
tubes.  The  canaliculi  of  the  outermost  row  of  lacunae  of  any  Haversian  system  do  not 
usually  open  into  the  lacuna  of  an  adjoining  system,  but  bend  on  themselves,  and  open  into  the 
lacuna  from  which  they  sprang,  or  into  an  adjoining  lacuna  of  the  same  system.  These  are 
the  so-called  '  recurrent '  canaliculi  (Ranvier).  {Indicate  the  lacuna:  and  canaliatli  in  PI.  VI., 
Figs.  4  and  5.) 

L.   S.    OF    DENSE    DRY   BONE. 

EXAMINATION  fL).— The  Haversian  canals  are  cut  longitudinally  or  obliquely  (with 
transverse  or  oblique  branches  connecting  them),  and  they  open  on  the  surface  of  the  bone 

F 


34  PRACTICAL   HISTOLOGY. 

and   into  the  medullary  cavity.     The   lacunae  are   lony,  narrow  ovals,  parallel  to   the   canals 
arranged  in  several  rows  between  adjoining  canals. 

BLOOD-VESSELS    OF    BONE. 

PREPARATION. — Place  the  leg-bones  of  a  rabbit  whose  blood-vessels  have  been  injected 
with  a  gelatine  and  carmine  mass  (p.  li),  in  a  three  per  cent,  solution  of  hydrochloric  acid 
for  several  days,  until  they  are  completely  decalcified.  Place  them  in  water  for  several  hours, 
and  afterwards  in  a  dilute  solution  of  sodic  carbonate,  to  get  rid  of  the  last  trace  of  acid. 
Make  longitudinal  and  transverse  sections  in  the  ordinary  way,  and  mount  them  in  Farrant's 
solution. 

EXAMINATION  (L). — Observe  each  Haversian  canal  contains  a  blood-vessel,  and  note  the 
general  arrangement,  which  follows  exactly  the  Haversian  canals.  {Indicate  the  longitudinal 
arrangement  of  the  blood-i'csscls  in  PI.  VI.,  Fig.  3.) 

CANCELLOUS    DECALCIFIED    RONE. 

PREPARATION. — Soften  the  head  of  a  foetal  bone  of  a  kitten  or  new-born  child  in  the 
usual  way,  and  make  transverse  sections  through  the  cancellous  head  of  the  bone.  Stain  a 
section  with  logwood  for  five  minutes,  and  another  with  picrocarmine  for  fifteen  minutes,  and 
mount  both  in  Farrant's  solution.     Cover. 

EXAMINATION  (L). — Observe  the  open  meshwork  ;  the  meshes  are  the  cancelli,  bounded 
by  trabeculee  of  bone,  which  do  not  contain  Haversian  canals.  In  the  cancelli  are  osteoblasts 
and  marrow-cells. 

(H).  Study  the  osteoblasts,  and  search  for  multi-nucleated  cells  (myoplaques,  giant-cells,  or 
osteoclasts) — cells  several  times  larger  than  the  osteoblasts,  containing  many  nuclei  imbedded 
in  granular  protoplasm.  They  usually  lie  close  on  the  bone,  in  a  depression  which  they 
accurately  fill. 

DEVELOPMENT   OF    BONE    (INTRA-CARTILAGINOUS). 

PREPARATION. — Place  the  leg-bones  of  a  rabbit,  one  week  old,  or  those  of  a  kitten,  in 
picric  acid  solution,  or  in  the  chromic  acid  and  nitric  acid  fluid,  till  they  are  completely  softened. 
Make  a  longitudinal  vertical  section  through  the  cartilaginous  head  and  shaft  of  the  bone. 

EXAMINATION  (L).— Stain  a  section  with  picrocarmine.  Observe  the  head  of  the  bone 
separated  from  the  shaft  by  the  epiphysial  cartilage,  and  below  the  cartilage  the  line  of  ossifi- 
cation. In  the  epiphysial  cartilage  the  cells  are  arranged  to  a  certain  depth  above  the  line  of 
ossification  in  rows  parallel  to  the  long  axis  of  the  bone.  These  rows  of  cells  are  produced 
by  transverse  cleavage  of  cartilage-cells.  Below  the  line  of  ossification,  where  true  bone  com- 
mences, are  found  irregular  spaces — the  primary  medullary  spaces — which  are  bounded  by 
narrow  spicules  of  calcified  cartilage,  partially  covered  by  a  laj-er  of  bone.  In  these  spaces 
are  seen  osteoblasts,  young  marrow-cells,  and  blood-vessels. 

(H).  Observe  the  rows  of  cartilage-cells,  smaller  above  and  larger  below.  At  the  line  of 
ossification  a  cartilage-capsule  may  be  found,  opening  and  discharging  its  contents  into  a 
primary  medullary  space.  The  long  spicules  have  in  their  centre  the  remains  of  the  calcified 
cartilage,  and  on  them  is  deposited  bone,  which  is  now  stained  of  a  deep  red  colour,  and 
osteoblasts  may  be  seen  in  process  of  becoming  imbedded  in  this  newly  formed  bone,  to  form 
bone-corpuscles.     In   the  spaces  observe  the  osteoblasts,  and  here  and  there  multi-nucleated 


OSSEOUS   TISSUE.  35 

cells  (osteoclasts),  lying  on    and  eroding  the   newly  formed   bone  of  the   osseous  trabeculae. 
{Indicate  the  rows  of  cartilage-cells,  line  of  ossification,  and  other  details,  in  PI.  VI.,  Fig.  7.) 

L.  S.    PHALANX    01"   A   FINGER   FROM    A   HUMAN    FCETUS. 

After  softening,  stain  with  picrocarmine  for  fifteen  minutes,  and  mount  in  Farrant's 
solution. 

EXAMINATION  (L). — Observe  the  shape  of  the  bone  under  the  periosteum  :  on  each  side 
of  the  shaft  is  a  triangular  piece  of  bone,  stained  bright  red  ;  the  base  of  each  triangular  por- 
tion corresponds  to  the  surface  of  the  shaft,  the  apices  pointing  inwards.  In  this  piece  of 
bone  are  seen  channels,  into  which  vessels  from  the  periosteum  proceed.  These  vessels 
are  accompanied  by  osteoblasts  into  the  interior  of  the  bone,  where  already  the  central 
part  has  been  excavated  to  form  primary  meduUaiy  spaces.  Towards  the  head  of  the  bone, 
the  cartilage-cells,  arranged  in  rows,  are  seen  just  as  in  an  epiphysial  cartilage.  Under  the 
perrosteum  there  are  one  or  more  layers  of  osteoblasts. 

DEVELOPMENT   OF    BONE   (INTRA-MEMBRANOUS). 

One  of  the  three  flat  bones  of  the  skull  of  a  fcetal  kitten  or  other  animal  is  taken,  and 
after  being  hardened  in  picric  acid  it  is  stained  with  picrocarmine  and  mounted  in  Farrant's 
solution. 

MEDULLARY    TISSUE   AND    MARROW. 

Ordinary  yellow  marrow  consists  chiefly  of  fat  (ninety  per  cent.),  but  the  spongy  ends  of 
bones  generally  and  the  medullary  cavity  of  the  long  bones  of  some  animals — e.g.  guinea-pig — 
contain  red  marrow. 

PREPARATION. — Take  the  leg-bone  of  a  rabbit  or  guinea-pig,  and  with  an  axe  or  chisel 
cleave  it  longitudinally.  With  the  point  of  a  knife  take  out  a  little  of  the  red  marrow,  and 
diffuse  it  in  a  drop  of  salt  solution  on  a  slide,  and  cover. 

EXAMINATION  (H). — Observe  the  marrow-cells  of  various  shapes — many  of  them  re- 
sembling colourless  blood-corpuscles,  though  tiiey  contain  a  large  clear  nucleus  ;  others  are 
identical  with  colourless  blood-corpuscles.  Large  multi-nucleated  cells,  many  times  the  size  of 
a  coloured  blood-corpuscle,  may  be  seen.  They  are  the  myeloplaques,  osteoclasts,  or  giant- 
cells,  and  consist  of  a  granular  mass  of  protoplasm,  containing  a  large  number  of  nuclei  in 
their  interior.  Dilute  alcoliol  reveals  these  nuclei,  and  frequently  causes  a  small,  transparent, 
bulla-like  mass  to  appear  on  the  side  of  the  cell  (Stirling).  » 

A  permanent  preparation  of  red  marrow  may  be  made  by  placing  a  piece  of  a  long  bone  of 
a  guinea-pig,  opened,  as  described  above,  in  dilute  alcohol  for  two  days.  Then  put  a  little  of 
the  marrow  on  a  slide,  and  stain  it  wjth  picrocarmine  or  logwood,  and  mount  the  preparation 
in  glycerine. 

TAIL   OF   A    RAT    DOUBLY-STAINED. 

PREPARATION.  Gold  chloride  and  an  aniline  dye. — See  p.  xlvii.,  where  this  process  is 
full}-  described. 

EXAMINATION  (L  and  H). — The  tendons  are  stained  with  gold  chloride,  while  the  bone 
is  stained  of  one  colour  and  the  cartilage  of  another.  This  is  one  of  the  most  beautiful  pre- 
parations in  the  whole  range  of  histology. 


36  PRACTICAL   HISTOLOGY 


MUSCULAR   TISSUE. 

There  are  two  varieties  of  muscular  tissue,  the  non-striped  and  the  striped.  The  former 
may  easily  be  obtained  from  certain  of  the  hollow  viscera,  and  the  latter  from  the  voluntary 
muscles  of  the  limbs. 

NOX-STRIPED    MUSCLE    (INTESTINE). 

PREPARATION.— Kill  a  rabbit  ;  wash  out  a  piece  of  the  small  intestine  with  salt  solution, 
and  afterwards  distend  the  gut  with  absolute  alcohol.  Place  the  distended  gut  in  absolute 
alcohol  for  twelve  hours.  After  it  is  hardened,  whilst  still  distended,  with  a  pair  of  forceps  strip 
off  a  thin  layer  of  the  longitudinally  disposed  muscular  fibres,  forming  the  outer  muscular  layer 
of  the  gut,  and  place  it  in  water  to  remove  the  alcohol  ;  float  it  on  to  a  slide  and  stain  with 
logwood  for  five  minutes  and  mount  it  in  dammar.     Cover. 

EXAMINATION  (L). — Observe  the  large  number  of  fusiform  nuclei  arranged  more  or  less 
parallel  to  each  other. 

(H).  Observe  the  fusiform  shape  of  the  nuclei  stained  deep  blue.  These  are  the  nuclei 
of  the  fusiform  muscular  fibre-cells  of  which  the  tissue  is  composed.  {^Indicate  the  uitdei  in 
PI.  VII.,  Fig.  I.) 

It  is  necessary  to  isolate  these  fusiform  cells  to  show  that  the  tissue  is  composed  of  a 
number  of  them  cemented  together. 

PREPARATION. — Place  a  small  piece  of  the  fresh  muscular  wall  of  the  intestine  or  stomach 
in  a  twenty  per  cent,  solution  of  nitric  acid  for  twenty-four  hours  ;  this  will  dissolve  the  cement. 
Place  a  strip  of  this  softened  tissue  in  water  to  remove  the  acid,  and  then  tease  a  small  piece 
on  a  slide.  It  is  very  difficult  to  stain  the  nuclei  of  these  isolated  cells  after  the  action 
of  nitric  acid,  but  a  watery  solution  of  magenta  may  be  tried  ;  or  steeping  in  picrocarmine  for 
forty-eight  hours  or  longer.     Mount  in  glycerine  and  cover. 

EXAMINATION  (H).— Notice  a  great  number  of  elongated  small  fusiform  cells  floating  in 
the  field.  Each  cell  is  broad  at  the  middle  and  tapers  towards  the  extremities.  Near  the 
centre  is  the  fusiform  nucleus  stained  red,  and  at  each  of  its  poles  not  unfrequently  a  few 
granules  are  to  be  seen.  In  some  of  the  fibres  corrugated  bars  may  be  seen  passing  across  the 
cell  ;  these  are  where  partial  contractions  of  the  body  of  the  cell  have  been  fi.xed  by  the  acid. 
{Indicate  these  fibres  in  PI.  VII.,  Fig.  2.) 

BLADDER   OF   A    FROG. 

PREPARATION.— Distend  the  bladder  of  a  frog  with  dilute  alcohol  (p.  xxxiv),  and  place  it 
distended  in  a  large  quantity  of  the  same  fluid  for  twenty-four  hours.  At  the  end  of  that  time 
slit  open  the  bladder,  place  it  on  a  slide  with  the  inner  surface  uppermost,  and  with  a  camel- 
hair  pencil  brush  away  all  the  epithelium  lining  the  bladder,  which  is  very  easily  removed. 
Wash  the  tissue  in  water,  and  stain  one  piece  with  logwood  and  mount  it  in  dammar,  and 
another  for  half-an-hour  with  picrocarmine,  and  mount  it  in  Farrant's  solution. 


MUSCULAR    TISSUE. 


o/ 


EXAMINATION  (L). —  Observe  the  large  number  of  nuclei  stained  either  blue  or  red,  lying 
usually  in  one  plane. 

(H).  Observe  the  shape  and  disposition  of  the  long  fusiform  nuclei.  Many  of  the  cells 
are  fusiform,  but  others  are  tri-radiate. 

T.  S.    OF    NON-STRIPED    MUSCLE. 

PREPABATION. — Place  a  piece  of  the  muscular  wall  of  the  intestine  of  a  cat  in  chromic 
acid  and  spirit  solution,  and  after  two  weeks  make  transverse  sections  of  its  circular  muscular 
fibres.  Stain  a  section  with  picrocarmine  for  twenty-four  hours  and  mount  it  in  Farrant's 
solution.     Stain  another  with  logwood  for  five  minutes,  and  mount  it  in  dammar. 

EXAMINATION  (H). — Observe  the  field  mapped  out  into  small  polygonal  areas  of  different 
sizes.  These  are  the  fibres  cut  across.  These  areas  occur  in  groups  of  small  fasciculi,  sepa- 
rated from  each  other  by  connective  tissue,  which  forms  a  sheath  for  them  —  the  periniysiuin. 
In  some  a  nucleus  may  be  found,  in  others  none,  because  the  line  of  section  has  not  been 
through  the  situation  of  the  nucleus  in  the  fibre.  The  variable  size  of  the  areas,  and  the 
absence  of  a  nucleus  in  some  of  them,  enable  the  observer  to  distinguish  easily  between  the 
cut  ends  of  non-striped  muscular  fibres  and  the  mosaic  formed  by  the  ends  of  a  number  of 
columnar  epithelium  cells  turned  towards  the  observer,  all  of  which  are  of  the  same  size,  and 
each  one  is  nucleated.  Observe  the  cement-substance  between  the  fibres.  This  is  best  seen 
when  the  illumination  is  slightly  diminished  by  turning  off  a  little  of  the  light  with  the 
mirror.  Observe  also  that  the  fibres  are  arranged  in  groups  or  bundles,  surrounded  and 
separated  from  each  other  by  connective-tissue  septa,  which  are  continuous  throughout — the 
periinysiuin.  This  is  continuous  with  the  cement  or  endoniysiiint,  which  lies  within  the  bundles 
between  the  muscle-cells.     {Indicate  the  divided  fibres  in  PI.  VII.,  Fig.  3.) 

CEMENT-SUBSTANCE    OF    NON-STRIPED    MUSCLE. 

PREPARATION. — Kill  a  rabbit,  and  wash  out  a  piece  of  the  small  intestine  with  distilled 
water.  Fill  the  intestine  with  a  half  per  cent,  solution  of  nitrate  of  silver,  and  place  the 
distended  intestine  for  ten  minutes  in  a  quarter  per  cent,  solution  of  nitrate  of  silver. 
Owing  to  the  difference  of  concentration  of  the  fluids  on  the  two  sides  of  the  gut,  diffusion  of 
the  silver  takes  place  and  so  complete  saturation  of  the  tissues  is  obtained.  Wash  away  the 
silver  in  ordinary  water  and  e.vpose  the  gut  in  alcohol  to  a  strong  light.  After  it  has  become 
brown,  with  a  pair  of  forceps  tear  off  a  thin  lamina  of  the  outer  longitudinal  muscular  wall  of 
the  gut ;  steep  it  in  water  and  float  it  on  to  a  slide  and  mount  it  in  Farrant's  solution.  Another 
strip  may  be  stained  with  logwood  solution  and  mounted  in  dammar,  if  it  be  desired  to  reveal  the 
nuclei  of  the  muscle-cells.  Another  method  of  obtaining  the  outer  layer  of  the  muscular  coat 
is  to  place  a  small  piece  of  the  gut  with  its  external  surface  next  the  glass  slide  ;  and  then 
with  a  knife  to  scrape  away  layer  after  layer  from  within  outwards  until  only  the  longitudinal 
and  the  serous  coatsTemain. 

EXAMINATION  (H). — Obsene  a  large  number  of  fine  black  lines  disposed  for  the  most 
part  longitudinally.  When  carefully  traced  they  are  seen  to  bound  narrow  fusiform  spaces  or 
areas.  These  silver  lines  indicate  the  existence  of  a  cement-substance  which  holds  the  fibres 
together.  Lying  between  the  fibres  are  elongated  channels  with  bulgings  upon  them,  and 
lined  by  sinuous  epithelium.  These  are  the  silvered  lymphatics,  i.e.  lacteals.  We  shall 
use  this  preparation  for  the  study  of  the  general  characters  of  lymphatics.  {Indicate  the  silver 
lines  in  PI.  VII.,  Fig.  4.) 


38  PRACTICAL   HISTOLOGY. 

INTRA-NUCLEAR   PLEXUS    OF    FIBRILS    IN    NON-STRIFEI)    MUSCLE. 
The  student  who  desires  to  show  this  arrangement  must  proceed  as  follows  : — 

PREPARATION. — Kill  a  newt  and  place  the  small  intestine  and  mesentery  in  a  five  per 
cent,  solution  of  ammonium  chromate  for  twenty-four  hours.  Wash  in  water  till  no  colour  is 
given  off,  then  transfer  it  to  picrocarmine  for  several  hours.  With  scissors  snip  off  a  small 
piece  of  the  mesentery  and  mount  it  in  glycerine.     Cover. 

EXAMINATION  (H). — Observe  the  very  large  spindle-cells,  either  single  or  disposed  in 
groups.  Study  a  nucleus  and  observe  in  its  interior  a  delicate  plexus  of  fibrils.  In  its  in- 
terstices lies  a  clear  homogeneous  substance.  On  carefully  focussing  with  one-eighth  of  an 
inch  objective,  fibrils  may  be  traced  through  the  poles  of  the  nucleus,  and  be  found  to  be  con- 
tinuous with  fibrils  disposed  in  the  long  axis  of  the  cell.  These  muscular  fibres  are  examined 
on  account  of  their  large  size  (Klein).      {Indicate  tlicsc  plexuses  of  fibrils  in  PI.  VII.,  Fig.  5.) 

MESENTERY    OF    NEWT    DOUBLY-STAINED    WITH    PICROCARMINE   AND    LOGWOOD. 

PREPARATION. — Prepare  the  mesentery  of  a  newt  with  ammonic  chromate  as  directed 
above.  After  washing  away  all  the  chromate  from  a  piece  of  the  mesentery,  stain  it  for 
fifteen  minutes  in  dilute  picrocarmine  (ten  drops  to  a  watch-glass  of  water).  Wash  it 
in  water  and  place  it  in  water  acidulated,  with  a  few  drops  of  acetic  or  picric  acid 
for  an  hour.  After  washing  place  it  in  dilute  logwood  solution,  until  it  assumes  a  faint  lilac 
colour,  taking  care  that  it  is  not  over-stained.     Mount  it  in  glycerine. 


STRIPED    MUSCLE. 

This  may  be  studied  in  any  of  the  voluntary  muscles.  A  muscle  consists  of  striped  muscu- 
lar fibres  held  together  by  connective  tissue  and  supplied  with  blood-vessels  and  nerves.  The 
fibres  must  be  examined  in  the  fresh  condition,  and  after  the  action  of  reagents. 

FRESH    STRIPED    MUSCLE. 

PREPARATION. — Kill  a  frog,  and  from  one  of  its  muscles — preferably  the  sartorius,  because 
it  is  composed  of  parallel  fibres — snip  off  a  small  piece  and  tease  it  slightly  in  salt  solution. 
Cover. 

EXAMINATION  (H), — Observe  the  cylindrical  fibres  consisting  of  the  sarcous  substance 
marked  with  alternate  light  and  dim  stripes  (PL  VII.,  Fig.  8).  In  some  places  the  striation 
may  be  obscure  or  the  stripes  very  close,  while  here  and  there  slight  longitudinal  fibrillation 
is  obser\'able.  Irrigate  with  distilled  water  and  after  a  short  time  a  fine  bleb  may  be  seen  on 
the  side  of  one  or  more  of  the  fibres.  This  is  the  sarcolemnia.  {Indicate  the  sarcolevnna  in 
PI.  VII.,  Fig.  6.)  Water  has  passed  through  the  sarcolemma  and  raised  it  from  the  sarcous 
substance.  Irrigate  the  preparation  with  dilute  acetic  acid,  when  the  nuclei,  lying  under 
the  sarcolemma  and  in  the  sarcous  substance,  are  revealed  (PI.  VII.,  Fig.  7).  They  are 
elongated  spindles,  sometimes  slightly  coloured,  lying  in  the  long  axis  of  the  fibres.  The 
reason  why  the  nuclei  are  not  seen  at  first  is  because  they  have  the  same  refractive  index  as 


MUSCULAR    TISSUE.  39 

the  surrounding  sarcous  substance.  Another  way  of  showing  the  sarcolemma  is  to  take  a 
piece  of  fresh  muscle,  and  after  teasing  it,  to  press  a  needle  across  a  fibre  so  as  to  try  and 
break  the  fibre.  If  this  be  accomplished,  the  sarcous  substance  will  be  ruptured,  and  con- 
tracting leaves  the  structureless  transparent  sarcolemma  stretched  between  the  ends  of  the 
ruptured  fibre. 

CLEAVAGE  OF  THE  SARCOUS  SUBSTANCE. 

PREPARATION. — Place  small,  narrow  strips  of  any  muscle  of  a  cat  or  rabbit,  which  has 
been  dead  for  a  few  hours,  in  a  small  quantity  of  a  sixth  per  cent,  solution  of  chromic  acid  for 
a  week.  The  muscular  fibres  will  tend  to  split  up  longitudinally  mtojibrils.  Chromic  acid 
seems  to  dissolve  a  cement. 

(2)  Place  similar  pieces  of  muscle  in  hydrochloric  acid — one  part  to  fifty  of  water  for  a 
week.  After  this  time  each  fibre  tends  to  split  transversely  into  discs.  Artificial  digestion 
(p.  92)  in  an  acid  medium  produces  the  same  result  in  a  few  hours. 

EXAMINATION  (H).— Take  a  small  piece  of  the  muscle  which  has  been  steeped  in 
chromic  acid,  and  after  washing  it  thoroughly  tease  it  out  with  needles  in  glycerine.  Each 
muscular  fibre  tends  to  split  longitudinally  into  excessively  fine  threads— the  fibrilhc  or 
fibrils — each  one  being  marked  v.'ith  light  and  dim  bands  like  the  original  fibre.  {Indicate  the 
fibnlhein  PL  VII.,  Fig.  10.) 

Take  a  similar  piece  of  the  hydrochloric  acid  muscle  and  tease  it.  The  fibres  split  across 
into  thin  narrow  discs  or  '  muscle-discs.'  Occasionally  a  muscular  fibre  splits  up  both  longitu- 
dinally and  transversely  at  the  same  time,  when  Bowman's  sarcous  elements  are  formed. 
These  are  sometimes  seen  at  the  end  of  a  fibre  broken  across  by  chance.  {Indicate  the  discs  in 
PL  VII.,  Fig.  9.) 

MUSCLES    OF   ARTHROPODA. 

To  trace  the  finer  details  in  the  structure  of  muscle  it  is  customary  to  use  the  striped 
muscles  of  insects,  such  as  the  common  water-beetle— Dytiscus  marginalis  or  Hydrophyllus 
piceus — or  the  common  cockroach.  I  find,  however,  that  the  muscles  of  the  common  edible 
crab  answer  the  purpose  admirably,  and  are  far  better  suited  for  use  in  a  large  class. 

PREPARATION.— (rt)  Place  a  beetle  or  the  amputated  limb  of  a  crab  in  absolute  alcohol, 
and  if  possible  keep  some  of  the  muscles  on  the  stretch  when  so  doing.  Leave  them  there  for 
a  week,  {b)  A  mixture  of  one  part  of  methylated  spirit  and  three  parts  of  Mailer's  fluid,  for 
three  to  four  weeks,  hardens  muscle  most  admirably.  The  tissue  must  be  kept  in  the  dark,  else 
reduction  of  the  chromic  salts  takes  place  very  rapidly.  Scoop  out  a  small  piece  of  the 
muscle  of  a  crab  and  steep  it  in  water.  Steep  one  piece  in  picrocarmine  for  twenty-four 
hours,  and  stain  another  with  logwood  for  a  few  minutes.  Tease  them  thoroughly  with 
needles.  Mount  the  former  in  glycerine  and  the  latter  in  dammar.  Both  preparations  show 
much  the  same  structure,  only  what  is  red  in  the  one  is  logwood-tinted  in  the  other. 

EXAMINATION  of  the  picrocarmine  preparation  (H). — Notice  the  large  size  of  the 
muscular  fibre,  and  how  easily  it  splits  into  fibrill^.  Each  fibre  shows  well-marked  transverse 
striation.  Study  this.  The  broad,  dim  discs  (contractile  discs)  are  stained  red,  and  the 
narrower,  less  refractive  discs  (interstitial  discs)  are  slightly  yellow.  In  each  dim  disc  a  series 
of  fine  vertical  rods  may  be  seen.  Study  carefully  the  light  disc — across  its  centre  runs  a 
fine  line,  Dobie's  line  or  Krause's  membrane — which  divides  the  disc  into  two  lateral  discs.  This 
line  is  continuous  with  the  sarcolemma,  and  is  supposed  to  represent  a  membrane  by  whicli 


40  PRACTICAL   HISTOLOGY. 

the  musclc-fibrc  is  divided  into  a  series  of  compartments,  each  containing  a  dim  disc  with  a 
lateral  disc  attached  to  each  end.  In  this  preparation  no  nuclei  are  seen.  {^Indicate  these  discs 
in  PI.  VII.,  Fig.  II.) 

Logwood  and  Lammar  Preparation  (H). — The  dim  discs  are  logwood-coloured — the  light 
discs  unaffected.  In  the  course  of  the  fibre  are  seen  oval  nuclei  under  the  sarcolemma. 
Otherwise  the  details  arc  the  same  as  in  the  above  preparation. 

If  the  muscle  of  an  insect— say  hydrophyllus  or  dytiscus  — be  taken  and  similarly  treated,  a 
row  of  fine  granules  is  observable  in  the  lateral  discs.  These  form  the  granular  layer  of 
Flogel. 

LIVING    MUSCULAR    FIBRE. 

PREPARATION. — This  is  not  particularly  well  adapted  for  a  large  class  of  students.  Cut 
off  the  head  of  a  water-beetle  and  then  open  its  chitinous  body  with  a  sharp-pointed  pair  of 
scissors,  so  as  to  expose  the  large  wing-muscles.  Snip  off  a  small  piece  of  these  and  tease  it 
rapidly  in  aqueous  humour  (p.  xxx)  or  salt  solution  (p.  xxx).     Cover  and  examine. 

EXAMINATION  (H). — Observe  the  fibres,  their  striation,  and  that  they  are  soft  and  plastic. 
Here  and  there  a  thickening,  progressing  along  a  fibre  — a  contraction-wave— with  approxi- 
mation of  the  discs,  may  be  observed. 


NUCLEI    OF    MUSCLE. 

PREPARATION. — Take  a  small  piece  of  a  muscle  of  a  rabbit  or  cat  which  has  been  pre- 
served in  alcohol,  tease  it  out  in  water,  and  stain  it  in  carmine  till  it  is  of  a  bright  red 
colour.  Wash  it  in  water  to  remove  the  surplus  carmine.  Place  on  it  a  large  drop  of  glacial 
acetic  acid,  and  after  doing  so  examine  it  with  a  low  power,  to  notice  when  the  nuclei  alone 
remain  stained.  The  effect  of  the  acid  is  to  remove  the  diffuse  staining,  and  leave  the  carmine 
only  in  the  nuclei.     Mount  it  in  either  Farrant's  solution  or  dammar — preferably  the  latter. 

EXAMINATION  (H). — Observe  the  striation,  and  in  the  course  of  the  fibres  oval  nuclei 
with  their  long  axis  in  the  axis  of  the  fibres,  stained  deeply  red.  {Indicate  the  nitdei  in  PL  VII., 
Fig.  8.) 

Relation  of  the  Nuclei  to  the  Sarcolemma. — Take  a  limb-muscle — or  the  tongue — of  a  cat 
or  rabbit  or  other  animal,  previously  hardened  in  alcohol,  or  chromic  acid  and  alcohol  (p.  x.xxi), 
and  make  a  transverse  section  by  freezing  or  other  means.  Stain  it  for  a  few  minutes  with 
logwood  and  mount  it  in  dammar.  This  preparation  will  also  show  the  relation  of  the 
connective  tissue,  ox  perimysium,  to  the  muscular  fibres. 

EXAMINATION  (L). — Notice  the  connective-tissue — perimysium — of  a  light  blue  colour 
surrounding  groups  of  muscular  fibres  cut  transversely,  which  appear  as  small  polygonal  areas 
stained  light  blue.  {Indicate  the  connective  tissue,  and  the  cut  muscular  fibres  with  their  nuclei 
in  PL  VII.,  Fig.  12.) 

(H). — Observe  the  cut  ends  of  the  fibres,  each  more  or  less  polygonal,  through  mutual 
pressure  of  the  fibres  against  each  other.  Under  the  sarcolemma  are  seen  two  or  more  small 
logwood-stained  nuclei  in  each  fibre,  and  the  cut  end  of  the  sarcous  substance  appears  slightly 
dotted.  The  nuclei  are  often  situated  at  the  angles  of  the  polygon.  Trace  the  connective 
tissue  in  very  fine  strands  between  the  fibres — forming  the  endomysium — and,  it  may  be,  con- 
taining a  capillary  here  and  there.  In  a  transverse  section  of  a  frog's  muscle  nuclei  are  found 
to  exist,  not  only  under  the  sarcolemma,  but  also  throughout  the  sarcous  substance. 


PLAi'E   V'^  MUSCLE 


ItntBrn  Bros  itk 


MUSCULAR    TISSUE.  41 


BLOOD-VESSELS    OF    MUSCLE. 

PEEPAE.ATION. — Make  longitudinal  and  transverse  sections  of  any  muscle  of  a  cat  or 
other  animal  whose  blood-vessels  have  been  injected  with  a  carmine  or  Prussian  blue  and 
gelatine  mass  (p.  li).  The  sections  may  be  very  slightly  stained  in  logwood,  or  mounted 
unstained  in  dammar,  or  a  smal!  piece  of  the  muscle  may  be  simply  teased  and  mounted  in 
Farrant's  solution. 

EXAMINATION  of  a  longitudinal  section  (L).  Observe  the  capillaries  arranged  parallel  to 
the  long  axis  of  the  fibres  —one  capillary  between  each  two  fibres,  with  here  and  there  trans- 
verse branches  connecting  them.  They  lie  outside  the  muscular  fibres  and  amongst  the 
delicate  connective  tissue  which  supports  them.  {^Indicate  the  arrangevient  of  the  capillaries  in 
PI.  VII.,  Fig.  13.) 

Transverse  section  (L). — Observe  the  cut  ends  of  the  fibres  and  between  and  outside  the 
sarcolcmma  the  cut  ends  of  the  capillaries.  The  larger  blood-vessels  lie  in  the  connective  tissue 
of  the  perimysium. 

It  is  advisable  to  inject  the  blood-vessels  of  the  lower  limbs  of  a  rabbit  with  a  Prussian 
blue  and  gelatine  mass.  Place  the  limbs  in  a  one  per  cent,  solution  of  bichromate  of  potash. 
After  the  mass  has  set,  select  one  of  the  red  muscles,  e.g.  the  semi-tendinosus.  Most  of  the 
muscles  of  the  rabbit  are  pale  or  almost  colourless.  The  above-mentioned  muscles,  and  some 
others  (soleus,  adductor  minus,  muscles  of  the  jaw),  are,  however,  distinctly  red  ;  the  semi-tendi- 
nosus, at  least,  has  a  peculiar  distribution  of  its  blood-vessels.  Tease  out  a  small  piece  of  one 
of  the  injected  red  muscles  and  mount  it  in  glycerine  or  dammar. 

EXAMINATION  (L). — The  blood-vessels  have  the  same  distribution  and  relations  as 
described  above,  but  they  present  this  peculiarity — that  on  the  transverse,  short  capillaries  are 
small  dilatations  like  little  aneurysmal  swellings.  These  swellings  do  not  exist  in  the  blood- 
vessels of  the  pale  muscles  of  the  same  animal.  Further,  the  capillaries  usually  pursue  a 
more  tortuous  course,  and  the  veins,  also,  sometimes  present  similar  fusiform  swellings  in 
their  course. 

The  muscular  fibres  of  the  heart  will  be  examined  in  connection  with  that  organ  (p.  47); 
and  the  nerves  of  muscles  with  the  terminations  of  nerve-fibres  (p.  45). 

Foetal  developing  muscle  may  be  studied  in  any  embryo  after  hardening  in  Miiller's  fluid. 
It  is  characterised  by  the  great  number  of  its  nuclei,  the  smallness  of  the  fibres,  and  the  im- 
perfect striation.     In  fresh  fcetal  muscle  iodine  solution  reveals  the  presence  of  glycogen. 

REL.4TI0N    OF    MUSCLE    TO   TENDON. 

PREPARATION  {a).  Centrum  tendineum  of  the  diaphragm. — Cut  out  the  diaphragm  from  a 
newly  killed  rabbit  ;  place  it  in  lemon  juice  for  five  minutes,  and,  after  steeping  in  one  per 
cent,  gold  chloride  for  an  hour,  transfer  to  a  twenty-five  per  cent,  solution  of  formic  acid  for 
twenty-four  hours.  During  this  time  the  preparation  must  be  kept  in  the  dark  till  the  gold 
is  reduced.  Snip  out  a  small  piece  of  the  muscle,  with  the  central  tendon  attached,  and 
tease  it  in  glycerine  on  a  slide  and  cover  ;  or  a  section  may  be  made  parallel  with  the  long 
axis  of  the  muscular  fibres. 

EXAMINATION  (Land  H). — Observe  the  tran.sversely  .striped  muscle,  and  trace  it  towards 
the  tendon,  where  it  ends  abruptly  in  a  conical  form  which  is  lodged  in  a  corresponding 
depression  in  the  tendon. 

G 


PRACTICAL   HISTOLOGY 


NERVE-FIBRES. 

There  are  two  varieties  of  nerve-fibres  : — 

1.  The  medullated  or  white. 

2.  The  non-medullated  or  gray. 

MEDULLATED  NERVE-FIBRES. 

Each  nerve-fibre  consists  of  a  thin  transparent  structureless  sheath,  the  primitive  sheath, 
with  here  and  there  a  nucleus  lying  under  it.  Within  this  is  the  medullary  sheath  or  white 
substance  of  Schwann  with  a  double  contour  forming  the  greater  part  of  the  nerve.  It  is 
interrupted  at  intervals  by  regular  constrictions,  or  Ranvier's  nodes,  where  the  white  substance 
is  entirely  absent.  In  the  centre  of  the  fibre  is  the  axis-cylinder,  a  fine  uniform  cylinder 
continuous  throughout  the  whole  length  of  the  fibre.  The  proper  view  to  take  of  the  axis- 
cylinder  is  that  it  is  directly  continuous  at  its  central  end  with  a  process  of  a  nerve-cell,  and 
is  in  fact  one  of  the  processes  of  a  nerve-cell  enormously  elongated  and  pushed  outwards 
towards  the  periphery  of  the  body,  and  covered  in  whole  or  part  of  its  course  with  one  or 
more  sheaths.  This  is  the  only  view  that  explains  the  remarkable  results  of  degeneration 
in  a  nerve-fibre  when  it  is  separated  from  its  so-called  trophic  nerve-centre  (Waller)  and 
also  explains  the  so-called  union  of  nerve-fibres,  which  is  in  fact  no  union  at  all,  but  is  due  to  a 
pushing  out  or  growth  of  the  axial  cylinder  into  the  old  sheaths  of  the  degenerated  nerve-fibres 
from  which  the  axis-cylinder  and  most  of  the  white  substance  of  Schwann,  but  not  the  nuclei, 
have  disappeared. 

FRESH   NERVE-FIBRES. 

PEEPAEATION. — Kill  a  frog  and  remove  its  sciatic  nerve.  With  a  pair  of  sharp  scissors 
cut  off  half  an  inch,  and  place  it  on  a  slide  in  a  drop  of  salt  solution.  With  a  pair  of  needles 
gently  tease  out  one  end  of  the  nerve,  and  examine  it  from  time  to  time  with  a  low  power,  to 
ascertain  when  it  has  been  sufficiently  teased.     Cover. 

EXAMINATION  (H). — Observe  the  nerve-fibres  of  various  sizes,  some  of  them  about  the 
breadth  of  a  coloured  blood-corpuscle,  others  larger  or  smaller.  Note  the  double  contour  of 
the  white  substance  of  Schwann  which  is  highly  refractive.  This  double  contour  exists  in  the 
living  nerve,  and  is  not  due  to  the  action  of  reagents.  This  can  easily  be  proved  by  examin- 
ing a  medullated  nerve  in  the  lung  or  tongue  of  a  living  frog  or  the  lung  of  a  newt.  Select 
the  end  of  a  fibre  which  has  been  broken  across,  and  study  the  mycline  exuding  from  its  cut 
end.  The  myeline,  or  white  substance,  collects  into  drops,  with  concentric  markings.  These 
must  not  be  mistaken  for  cells,  from  which  they  are  easily  distinguished  by  the  absence  of  a 
nucleus.     {Indicate  the  myeline  exuding  from  a  nerve-fibre  in  PI.  VIII.,  Fig.  i.) 

In  the  centre  of  each  fibre  is  a  clear  uniform  band,  the  axis-cylinder.  Trace  the  course  of 
a  fibre  carefully,  and  a  Ranvier's  node  may  be  easily  distinguished.  The  white  substance  is 
interrupted  here  ;  further,  if  the  nerve-fibre  be  slightly  on  the  stretch,  the  oblique  incisures  of 
Schmidt  and  Lantermann,  passing  through  the  myeline,  can  also  be  made  out.  Irrigate  the 
preparation  with  picrocarmine,  and  continue  the  examination  after  twenty  minutes.  Observe 
that  a  nucleus  under  the  primitive  sheath  is  stained  red,  the  carmine  passes  in  at  the  nodes  of 
Ranvier  and  also  stains  the  axis-cylinder  red.     If  an  axis-cylinder  happen  to  extend  beyond 


NERVE-TISSUE.  43 

the  myeline,  it   is   quickly  stained    red  by  the  carmine.     {Indicate  a  node  and  the  incisures  in 
PI.  VIII.,  Fig  2.) 

It  is  remarkable  that  nearly  all  the  phenomena  which  have  been  laboriously  made  out  by 
the  use  of  special  reagents  can  be  seen  in  a  perfectly  fresh  or  even  in  a  living  nerve-fibre. 

EFFECT  OF   OSMIC  ACID. 

PREPARATION. — Kill  a  frog  ;  dissect  out  a  sciatic  nerve.  Tie  a  fine  thread  round  each 
end  of  the  nerve,  carefully  preserving  it  from  getting  dry,  and  by  means  of  the  thread  stretch 
the  nerve  on  a  small  piece  of  wood— an  ordinary  wooden  match  with  a  slit  at  either  end  to 
hold  the  thread  answers  admirably.  Place  the  nerve  and  wood  in  a  one  per  cent,  osmic  acid 
solution  for  ten  minutes.  The  effect  of  the  osmic  acid  is  to  blacken  the  phosphorised  fats 
of  the  myeline  and  to  '  fix  '  its  elements.  The  nerve  is  stretched  in  order  that  the  nodes  and 
incisures  may  be  made  quite  distinct.  Remove  it  and  wash  it  carefully  in  water,  and  then 
place  it,  still  on  the  stretch,  in  a  test-tube  with  picrocarmine.  In  twenty-four  hours  the  stain- 
ing is  complete.  I  find  it  to  be  very  advantageous  to  leave  it  in  picrocarmine  for  a  fortnight 
or  longer.  The  effect  of  this  is  to  soften  the  connective-tissue  of  the  nerve,  so  that  on  teasing 
it  at  the  end  of  this  time  the  single  nerve-fibres  are  easily  obtained.  Tease  a  small  piece — 
always  tearing  off  a  fibre  in  its  long  axis — in  a  drop  of  glycerine.     Cover. 

EXAMINATION  (H). — Observe  the  myeline,  blackened  and  still  retaining  its  double  con- 
tour. Trace  a  fibre  and  observe,  at  pretty  regular  intervals,  the  constrictions  or  nodes  of 
Ranvier,  and  between  each  two  nodes  a  brightly  stained  nucleus  imbedded  in  a  trace  of  pro- 
toplasm, will  be  found  lying  under  the  primitive  sheath  and  partly  indenting  the  myeline. 
{Indicate  tii'o  nodes  in  PI.  VIII.,  Fig.  5.)  Study  a  node.  Observe  the  absence  of  the  myeline, 
and  that  only  the  slightly  red-coloured  axis-cylinder  passes  from  one  segment  of  the  nerve  to 
another.  The  axis-cylinder,  therefore,  is  the  only  essential  part  of  a  nerve.  Some  excessively 
delicate  nerve-fibres  will  be  seen.  The  delicate  connective  tissue  supporting  the  nerve-fibres 
is  easily  made  out.     {Indicate  a  node,  the  nucleus,  and  the  incisures  in  PI.  VIII.,  Figs.  3  and  4.) 

Amongst  the  medullated  fibres  may  be  detected  the  grey  or  sympathetic  nerve-fibres. 
They  are  distinguished  by  being  delicate  fibres,  with  a  large  oval  nucleus  here  and  there  in 
their  course,  and  are  devoid  of  myeline,  and  therefore  do  not  possess  a  double  contour. 

ACTION  OF  NITRATE  OF  SILVER.     RANVIER'S  CROSSES. 

PREPARATION. — {a)  Tease  out  roughly  a  small  piece  of  a  perfectly  fresh  sciatic  nerve  of 
a  frog  in  a  quarter  per  cent,  solution  of  silver  nitrate  on  a  slide,  and  leave  it  in  this  solution  for 
five  minutes  or  so.  Wash  it  with  water  and  then  tease  it  carefully  in  glycerine,  cover  and 
expose  it  to  the  action  of  light  until  it  becomes  brown. 

(3)  The  small  intercostal  nerves  of  a  rat  or  rabbit  may  be  excised  and  plunged  entire  into 
silver  nitrate  solution  and  mounted  entire  in  glycerine,  and  exposed  to  the  action  of  light  as 
above. 

EXAMINATION  (L  and  H). — Observe  a  fibre  and  notice  the  dark  brown  crosses  seen  here 
and  there  (PL  VIII.,  Fig.  8),  The  long  limb  of  the  cross  is  produced  by  the  silver  pene- 
trating to  the  axis-cylinder,  and  staining  it,  or  an  albuminous  material  covering  it,  for  a  short 
distance  ;  whilst  the  transverse,  more  deeply  stained,  bar  is  due  to  the  cement  which  unites  the 
so-called  segments  of  a  nerve  one  to  another  (PI.  VIII.,  Fig.  9).  Sometimes  the  longitudinal 
limb  of  the  cross  may  be  marked  transversely  bylines  (Frommann's  lines).  If  a  whole  nerve 
be  used,  a  complete  investment  of  endothelial  cells  will  be  found  outside  all. 


44  PRACTICAL    HISTOLOGY. 


ARRANGEMENT  OF  NERVE-FIBRES  TO  FORM  A  NERVE. 

PREPARATION. — Place  pieces  of  the  human  or  other  sciatic  nerve,  cut  into  pieces  one  inch 
long,  into  chromic  acid  and  spirit  (p.  xxxi),  or  Miiller's  fluid  and  spirit  (p.  xxxii),  for  ten  days,  and 
after  hardening  in  alcohol  make  transverse  sections  in  the  ordinary  way.  Stain  a  section  in 
logwood,  and,  if  it  be  desired,  another  in  carmine,  and  mount  them  in  dammar.  They  both 
show  the  same  details. 

EXAMINATION  (L). — The  various  bundles  composed  of  nerve-fibres  are  held  together  by  a 
common  framework  of  connective  tissue — the  epineurium — arranged  so  as  to  form  a  sheath 
around  and  between  the  various  bundles,  and  in  it  may  be  found  transverse  sections  of  the 
large  blood-vessels  which  supply  the  nerve  with  blood.  Round  each  bundle  observe  a  special 
sheath — \\\&  perincuruiiii — composed  of  lamellated  connective  tissue  with  flattened  connective- 
tissue  cells  between  them.  The  spaces  between  the  lamellae  represent  lymph-spaces.  {Indicate 
these  bundles  in  PL  VIII.,  P"ig.  lO.) 

Study  a  nerve-bundle. — Observe  it  is  made  up  of  many  nerves  cut  transversely  ;  only  the 
stained  axis-cylinders  are  indicated. 

(H).  Study  the  epi-  and  perineurium  ;  observe  the  cut  ends  of  the  nerve-fibres  varying  in 
diameter,  and  the  section  of  the  a.xis-cylinder  stained  red  or  violet  and  surrounded  by  a  trans- 
parent circular  area,  which  represents  the  myeline  or  white  sheath  of  Schwann  ;  outside  this 
a  faint  circle,  indicating  the  primitive  sheath.  Between  the  nerve-fibres  may  be  seen  very 
delicate  connective  tissue — the  endoneiiriuni — with  here  and  there  a  small  blood-vessel  in  it. 
{Indicate  the  stained  axial  cylinders  and  the  cut  ends  of  the  nerve-fibres  in  PL  VIII.,  Fig.  1 1.) 
If  the  hardening  process  has  been  continued  for  a  month  or  longer,  concentric  rings  may  be 
observed  in  the  white  substance  of  Schwann.  {Indicate  these  in  PL  VIII.,  Fig.  12.)  Longi- 
tudinal sections  of  a  nerve  prepared  as  above  may  also  be  studied. 

THE  CONNECTIVE-TISSUE  ELEMENTS  OF  A  NERVE. 

PREPARATION. — Take  a  small  fragment  of  a  nerve  which  has  been  hardened  in  one-si.xth 
per  cent,  solution  of  chromic  acid  for  ten  days,  and  after  staining  in  logwood,  tease  it 
thoroughly  in  a  drop  of  glycerine. 

EXAMINATION  (H). — Observe  the  nerve-fibres.  The  axis-cylinder,  stained  violet,  will 
easily  be  recognised  amidst  the  coagulated  myeline.  It  is  specially  interesting  to  trace  the 
axis-cylinder  across  a  node  of  Ranvier.  Observe  the  connective  tissue,  some  of  it  in  the  form 
of  delicate  lamellae,  with  here  and  there  oval  nuclei  stained  violet  lying  on  them,  and  also  de- 
tached flattened  plates— the  connective-tissue  corpuscles — each  containing  a  large  oval  violet 
nucleus. 

FRESH  NERVE-FIBRES  OF  THE  SPINAL  CORD. 

PREPARATION. — With  scissors  snip  off  a  small  fragment  of  the  white  matter  of  a  fresh 
spinal  cord  of  an  o.x  or  sheep,  place  it  on  a  dry  slide,  do  not  add  any  fluid,  put  on  a  cover- 
glass,  and  compress  it  into  a  thin  layer. 

EXAMINATION  (H).— Observe  the  drops  of  myeline,  easily  recognised  by  their  concentric 
markings  and  the  absence  of  a  nucleus,  and  very  delicate  nerve-fibres  with  bulgings  or  am- 
pulla: upon   them,  due   to  the   pressure  of  the  cover-glass  on  the  nerve-fibres,  which  here  are 


y.  300 


L  ; 


Plate  vm  ^  Nerves 


12 


11 


V.  -> 

n 


8, 


ilmtem  Bros  lith 


NERVE-TISSUE.  45 

devoid  of  a   primitive   sheath.     There  are   no   ampullae  in  the  living  condition.     Do  not  pre- 
.serve  this  preparation. 

NON-MEDULLATEU    NERVE-FIBRES. 

These  may  be  detected  in  an  ordinary  nerve  and  studied  there,  but  it  is  better  to  select  a 
nerve  containing  these  or  sympathetic  fibres  in  large  quantities.  The  sympathetic  nerve  from  the 
neck  of  a  rabbit  may  be  selected,  but  it  is  far  better  to  have  a  larger  nerve.  This  is  easily 
obtained  from  the  spleen  of  an  ox.  Accompanying  the  branches  of  the  splenic  vein  are 
large  nerve-trunks,  the  nerves  of  the  spleen,  which  contain  only  a  few  meduUated  fibres,  the 
rest  being  non-medullated  nerve-fibres.  With  scissors  cut  open  the  splenic  vein,  and  then  the 
nerve-trunk  is  easily  dissected  out. 

PREPARATION. — Place  the  nerve,  extended,  as  for  the  medullated  nerve,  in  a  one  per 
cent,  solution  of  osmic  acid,  and  after  washing  place  it  in  picrocarmine  for  twenty-four  hours. 
Tease  a  small  fragment  in  glycerine,  and  cover. 

EXAMINATION  (H). — Neglect  the  medullated  nerve-fibres  and  observe  the  non-medullated 
nerve-fibres.  They  are  pale,  flat,  slightly  granular  bands  which  have  no  myeline,  and  in  their 
course  the  oval  red-stained  nuclei  of  the  primitive  sheath  are  easily  seen.  Sometimes  the  nuclei 
are  seen  on  edge,  and  at  other  times  on  the  flat.  In  this  nerve  many  '  plasma  cells  '  (p.  20) 
are  found. 

If  desired,  a  fresh  nerve  may  be  teased  in  salt  solution,  when  similar  appearances  are 
observable. 

Nerve-cells  will  be  considered  with  the  organs  in  which  they  occur. 

SOME   PERIPHERAL  TERMINATIONS  OF    NERVES. 

We  shall  only  allude  here  in  detail  to  the  terminations  of  nerves  in  striped  muscle  by 
means  of  end-plates,  and  to  the  Pacinian  corpuscles  attached  to  the  ends  of  certain  sensory 
nerves  (p.  46).     Other  modes  of  termination  will  be  described  in  the  organs  where  they  occur. 

TERMIN-AFIONS  IN   STRIPED  MUSCLE. 

PREPARATION. — This  is  by  no  means  an  easy  task.  In  mammals  the  best  muscles  are 
the  straight  muscles  of  the  eyeball  of  a  rabbit — the  muscular  part  of  the  diaphragm  and  any 
thin  muscles  such  as  the  intercostals.  In  the  lizard  {Lacerta  agilis)  the  end  plates  may  be 
more  easily  found.  But  the  best  muscles  are  those  which  attach  the  skin  to  the  ribs  of  the 
smaller  snakes.  In  any  case  it  is  best  to  employ  the  following  gold  method.  Snip  out  very 
thin  pieces  of  the  muscle  to  be  examined,  and  place  them  in  lemon-juice  for  five  minutes,  then 
wash  in  water  and  transfer  to  a  one  per  cent,  solution  of  gold  chloride  for  twenty  minutes  or  half 
an  hour — wash  again  and  place  the  prepai-ation  in  the  dark  for  twenty-four  hours  in  a  twenty- 
five  per  cent,  solution  of  formic  acid.  After  washing  thoroughly,  to  remove  the  acid,  tease 
out  a  small  piece  of  the  muscle  in  glycerine,  and  search  with  a  low  power  for  a  nerve-fibre,  or 
trace  the  outline  of  a  muscular  fibre  till  an  end  plate  is  obtained.  The  a.xis-cylinder  is  con- 
tinued beyond  the  end  plate,  and  breaks  up  into  fibrils  inside  the  sarcolemma  ;  their  ulti- 
mate relation  to  the  sarcous  substance,  however,  is  unknown.  These  preparations  are  difficult 
to  make,  and  do  not  keep  well  afterwards. 

The  terminations  of  nerves  in  unstriped  muscle  will  be  considered  under  the  head  of  small 
intestine  (p.  70  ;  the  same  methods  are  applicable  as  for  those  of  striped  muscle  nerve-endings. 


46  PRACTICAL   HISTOLOGY. 


PACINI'S  CORPUSCLES. 

PREPARATIOIf. — These  are  numerous  in  the  mescnter)'  and  in  the  mcsorectum  of  the  cat, 
where  they  arc  easily  obtained  free  from  fat.  The  student  may  also  find  them  in  the  deeper 
layers  of  a  vertical  section  of  the  skin  of  the  hand  or  foot,  or  in  the  pad  of  a  cat's  foot,  but  I 
have  found  them  most  easily  for  students  in  transverse  sections  of  an  entire  fcetal  leg  or 
arm.  Being  placed  relatively  closer  to  each  other  in  the  fcetus,  they  are,  like  sweat-glands, 
more  easily  observed  there  than  in  adult  textures. 

In  the  cat's  mesentery  they  can  be  seen  as  small,  oval,  transparent,  hard  bodies.  Snip  out 
one  of  these  and  examine  it  at  once  in  salt  solution  with  a  low  power  ;  the  nerve  entering  it 
may  be  seen,  as  also  the  central  core  surrounded  by  concentric  laminae.  If  a  permanent  pre- 
paration be  desired,  excise  a  piece  of  the  mesorectum  or  mesentery  containing  these  bodies, 
pin  it  out  in  a  stretched  condition  on  a  flat  piece  of  cork,  and  place  the  cork,  tissue  down- 
wards, for  a  week,  in  a  two  per  cent,  solution  of  potassic  bichromate  ;  after  that  wash  away  all 
trace  of  the  reagent  with  water,  snip  out  a  corpuscle  and  stain  it  slowly  (twenty-four  to  forty-eight 
hours)  in  dilute  logwood,  which  must  be  changed  several  times.  Wash  in  water  and  mount  in 
glycerine.  As  the  corpuscles  are  of  considerable  thickness,  it  may  be  necessary  to  place  a  thin 
layer  of  paper  under  the  edges  of  the  cover-glass  to  avert  pressure,  and  allow  the  cover-glass 
to  lie  flat  upon  the  preparation. 

EXAMINATION  (L). — Observe  the  nerve-fibre  like  a  stalk  approaching  and  entering  the 
central  core  of  the  oval  corpuscle,  which  latter  is  made  up  of  laminse  one  outside  the  other. 
(H).  Trace  the  nerve-fibre,  and  observe  that  the  medullary  sheath  stops  where  the  nerve 
enters  the  corpuscle.  Only  the  axial  cylinder  passes  into  the  central  core.  Study  the  con- 
centric laminae,  which  exhibit  a  series  of  nuclei  on  each  surface.  These  are  the  nuclei  of  the 
endothelial  cells  which  cover  each  lamina. 

To  study  the  laminae  and  their  endothelial  covering,  a  corpuscle,  hardened  as  above,  must 
be  broken  up  with  needles,  by  which  process  the  various  lamina;  are  readily  separated  from 
each  other. 

Transverse  sections  will  be  obtained  in  one  or  other  of  the  preparations  already  mentioned. 
These  show  the  concentric  distribution  of  the  laminae  round  the  central  core,  and  also  the 
nuclei  of  the  cells  between  each  two  laminae  (p.  92). 


47 


HEART  AND  BLOOD- VESSELS. 

The  heart  is  invested  by  a  fibrous  pericardium  covered  by  a  layer  of  squames,  whose 
existence  is  easily  demonstrated  by  the  nitrate  of  silver  process.  It  is  lined  by  a  thinner 
membrane  also  covered  with  squames,  the  endocardium,  capable  of  demonstration  in  a  similar 
manner.  The  muscular  tissue  of  the  heart  is  transversely  striped,  though  involuntary.  The 
strise  are  always  more  or  less  indistinct.  The  fibres  branch  and  anastomose.  They  consist  of 
short  nucleated  segments  joined  by  a  cement,  and  are  devoid  of  a  sarcolemma. 

ISOL.'VTED  MUSCULAR  FIBRES  OF  THE  FROG'S  HEART. 

PREPARATION. — Place  the  heart  of  a  newly  killed  frog  in  a  small  quantity  of  a  forty  per 
cent,  solution  of  caustic  potash  (p.  xxxiv)  for  a  quarter  of  an  hour.  Take  a  fragment  of  the 
tissue  and  tease  it  with  needles  in  a  drop  of  potash  solution.  Be  careful  to  add  no  water. 
Cover. 

EXAMINATION  (H). — Observe  the  isolated  muscle-cells.  They  are  fusiform,  and  in  the 
broad  middle  part  there  is  a  well-defined  nucleus,  and  the  substance  of  the  fibre  is  trans- 
versely striped.  Do  not  preserve  this.  If  it  be  desired  to  possess  a  permanent  preparation, 
small  pieces  of  a  frog's  heart  must  be  placed  in  dilute  alcohol  for  twenty-four  or  forty  hours, 
and  then  stained  in  picrocarmine  ;  and,  after  teasing  to  isolate  individual  cells,  add  glycerine 
and  mount  the  preparation. 

FRESH    HEART   OF   A  MAMMAL. 
Treat  a  small  piece  of  the  heart  of  a  rabbit  in  the  same  way  as  the  frog's  heart.     Cover. 
EXAMINATION  (H).— Observe  the  oblong   muscle-cells  isolated.     Each  cell  is  nucleated 
and  transversely  striated.     Do  not  preserve  this. 

HEART    FOR   PRESERVATION. 

PREPARATION.— Place  small  pieces  of  the  heart  of  a  rabbit,  or  a  piece  of  a  human  heart, 
including  a  piece  of  the  pericardium,  in  chromic  acid  and  spirit  fluid  for  ten  days,  transfer  to 
spirit,  and  then  make  a  transverse  section  through  a  ventricle,  including  the  pericardium.  Stain 
a  section  with  logwood  and  mount  it  in  dammar. 

EXAMINATION  (L).— Observe  the  fibrous  pericardium  stained  of  a  light  blue,  and  note  its 
thickness.  Here  and  there  it  sends  fine  processes  into  the  heart,  to  form  a  perimysium  for  the 
bundles  of  muscle.  It  is  important  to  observe  the  normal  thickness  of  the  fibrous  covering, 
and  also  the  normal  amount  of  connective  tissue  present,  as  m  every  organ,  to  be  enabled  to 
judge  when  there  is  any  alteration  in  the  thickness  of  the  capsule  or  the  amount  of  the  inter- 
stitial connective  tissue.  This  point  is  specially  important  in  relation  to  the  pathological 
histology  of  organs.     Not  unfrequently  in   these  trabecule   are  to   be  seen   sections   of  the 


48  PRACTICAL    HISTOLOGY. 

coronary  artery  and  vein  and  cardiac  nerves,  easily  recogjnised  by  their  lying  together  and  by 
their  structure.  Trace  the  muscular  fibres  mapped  out  into  bundles.  Some  are  cut  trans- 
versely, others  obliquely,  and  some  longitudinally.  Many  nuclei  are  seen,  which  are  the  nuclei 
of  the  muscular  fibres.  Select  a  transverse  section  of  a  group  of  fibres.  (H).  Observe  the 
polygonal  shape  of  the  fibres  from  mutual  pressure,  the  existence  of  a  violet-stained  nucleus 
in  the  centre  of  some  of  them — not  in  all,  and  the  striae  radiating  from  the  nucleus  ;  a  delicate 
endomysium  will  also  be  seen.  {Indicate  these  in  PI.  IX.,  Fig.  2.)  Select  a  longitudinal  section, 
observe  the  faint  striation,  the  branching  and  anastomosing  of  the  fibres,  and  the  oval,  violet- 
stained  nuclei  in  the  course  of  the  fibres.  The  places  where  the  oblong  muscular  cells  which 
make  up  a  fibre  join,  indicated  by  a  transverse  line,  due  to  the  presence  of  a  cement-sub- 
stance, are  to  be  looked  for.     {Indicate  the  characters  of  these  fibres  in  PI.  IX.,  Fig.  i.) 

Cement-substance. — The  cement-substance  which  unites  one  cell  to  another  may  be  de- 
monstrated thus.  Thrust  the  nozzle  of  a  hypodermic  syringe  filled  with  half  per  cent,  solution 
of  silver  nitrate  amongst  the  fibres  <  T  ,-the  heart,  and  forcibly  inject  the  silver  solution  so  as 
to  separate  the  heart-fibres.  Place  the  her-t  in  alcohol,  and  after  twenty-four  hours  snip  out  a 
small  piece  which  has  been  acted  on  by  the  silver,  and  after  teasing  in  glycerine  expose  it  to 
the  light  and  examine  for  the  lines  of  junction  of  the  cells  stained  brown  or  black  by  the 
silver. 

Blood-vessels  of  the  heart  present  practically  the  same  arrangement  as  in  striped  muscle,  and 
are  prepared  and  examined  in  the  same  way  (p.  41). 

Nerves  and  ganglia  of  the  heart. — Sometimes  preparations  of  these  are  obtained  in  trans- 
verse .sections  of  the  ventricles  ;  but  to  see  their  arrangement  and  distribution  the  heart  of  a 
frog,  or  small  pieces  of  any  mammalian  heart,  must  be  treated  with  lemon-juice,  chloride  of 
gold,  and  formic  acid,  as  described  at  p.  xlv.  The  nerves  and  ganglia  must  then  be  dissected 
out  with  the  aid  of  a  dissecting  microscope  (p.  xxxv). 

PURKINJE'S    FIBRES. 

A  delicate  network  of  fine  transparent  lines  is  seen,  with  the  naked  eye,  on  the  interior 
of  the  ventricles  of  the  heart  of  a  sheep  or  ox.  These  are  Purkinje's  fibres.  They  consist  of 
imperfectly  developed  muscular  fibres — they  are,  in  fact,  vestiges  of  what  each  heart-fibre  was 
originally.  They  are  made  up  of  polygonal-shaped  cells,  each  often  with  two  nuclei,  and  only 
the  outer  half  of  each  cell  is  striated  ;  but  in  the  deeper  layers,  cells  are  found  where  the 
striation  has  involved  three- fourths  of  a  cell,  and  deeper  down  still  they  become  continuous 
with  the  ordinary  heart-muscle.  Each  muscular  fibre  of  the  heart,  and  in  fact,  every  striped 
muscle,  was  originally  non-striped,  and  the  striation  always  begins  at  the  periphery  of  the  cell, 
so  that  the  fibres  made  up  of  these  cells  represent  an  embryonal  phase  of  muscular  tissue. 

PREPARATION. — With  scissors  or  a  knife  cut  out  these  fine  threads  and  macerate  them 
for  two  days  in  dilute  alcohol,  stain  with  picrocarmine,  and  tease  a  small  piece  in  glycerine 
until  isolated  cells  or  groups  of  cells  are  found. 

EXAMINATION  (H). — Observe  the  isolated  polygonal  cells,  each  with  two  nuclei,  and 
the  outer  half  of  each  cell  marked  with  every  gradation  between  these  and  cells  where  the  peri- 
nuclear part  is  completely  striated.     {Indicate  these  cells  in  PI.  IX.,  Fig.  3.) 


49 


BLOOD-VESSELS. 

There  are  three  well-marked  varieties  of  blood-vessels — arteries,  veins,  and  capillaries.  A 
typical  artery  has  three  coats — outer,  middle,  and  inner.  In  a  middle-sized  artery  the  inner  coat 
consists  of  a  layer  of  elongated  squames  with  their  long  axis  corresjionding  to  the  long  axis  of 
the  vessel  ;  outside  this,  delicate  connective  tissue  and  an  elastic  membrane  or  lamina,  either 
with  holes  in  it — '  fenestrated  membrane  ' — or  it  consists  of  a  sheet  of  elastic  tissue  which  in 
sections  is  thrown  into  folds,  owing  to  the  contraction  of  the  muscular  coat  outside  it.  It  is 
to  be  remembered,  however,  that  these  folds  do  not  exist  in  a  living  vessel  distended  with 
blood.  The  middle  layer  consists  of  circularly-disposed  non-striped  muscular  fibres,  with  a 
small  amount  of  elastic  and  white  fibres.  The  outer  coat  (adventitia)  consists  of  fibrous 
tissue  with  longitudinally  disposed  elastic  fibres,  which  are  specially  numerous  next  the 
muscular  layer.  In  the  largest  arteries  (aorta)  there  is  relatively  a  large  amount  of  elastic 
tissue  in  the  inner  and  middle  coats,  and  in  the  middle  coat  the  non-striped  muscle 
and  elastic  plates  occur  in  alternate  layers.  In  the  veins  the  division  into  coats  is  not 
so  sharply  marked.  The  middle  coat  contains  less  muscular  fibre-cells  and  more  connective 
tissue,  and  the  outer  coat  not  unfrequently  contains  non-striped  muscle,  especially  in  large 
veins.  The  capillaries  consist  of  a  layer  of  nucleated,  flattened,  epithelial  cells,  joined  at  their 
edges  with  a  cement-substance. 

Small  arteries  and  capillaries  are  best  studied  in  the  pia  mater. 

SMALL   ARTERIES   AND    CAPILLARIES. 

PREPARATION. — Get  the  head  of  a  recently  killed  sheep,  and  remove  the  brain.  With  a 
scalpel  remove  as  much  of  the  brain  as  possible,  leaving  the  pia  mater.  With  a  stream  of 
salt  solution  wash  away  the  remainder  of  the  brain-substance.  It  is  most  important  to 
remove  all  the  brain-substance,  else  it  forms  a  granular  deposit  which  obscures  the  prepara- 
tion. Place  the  pia  mater  in  chromic  acid  and  spirit  solution  for  a  week.  After  washing  it 
thoroughly  in  water,  snip  out  a  small  piece  and  stain  it  with  logwood,  and  mount  it  in  dammar. 
A  similar  preparation  may  be  stained  with  picrocarmine,  and  mounted  in  Farrant's  solution. 

EXAMINATION  (L). — Select  a  small  artery,  an.d  study  its  mode  of  branching  and  how 
these  branches  terminate  in  capillaries.  Note  in  the  artery  the  transversely  disposed  nuclei  of 
the  muscular  fibres  of  its  middle  coat  (PI.  IX.,  Figs.  6  and  7).  Observe  the  outer  coat.  Search 
for  a  vein  with  its  much  thinner  coat  and  fewer  transversely  disposed  nuclei — i.e.  muscular 
fibres — and  compare  it  with  an  artery. 

(H).  Select  a  small  artery.  Observe  its  outer  and  middle  coats,  and,  composing  the  latter, 
non-striped  muscular  fibres,  disposed  in  one  or  more  layers.  The  fusiform  nuclei  of  the 
muscular  coat  are  very  apparent.  The  folds  of  the  elastic  lamina  of  the  inner  coat  may  be 
seen.  Study  a  capillary.  It  appears  as  a  perfectly  structureless  membrane,  with  oval  nuclei 
imbedded  in  it,  which  bulge  into  its  lumen.  It  requires  the  use  of  nitrate  of  silver  to  demon- 
strate its  endothelial  characters. 


H 


50  PRACTICAL   HISTOLOGY. 


DEVELOPMENT    OF    CAPILLARIES. 

The  development  of  capillaries  is  easily  studied  in  the  tail  of  a  half-grown  tadpole  prepared 
by  the  ordinary  gold  chloride  method.  To  see  the  capillaries  distinctly  it  is  well  to  remove 
the  superficial  epithelium  after  staining.  If  the  lemon-juice  and  gold  method  (p.  xlv)  be 
employed,  the  formic  acid  employed  afterwards  removes  the  superficial  epithelium.  The 
omentum  of  a  new-born  rabbit  presents  easily  recognised  whitish  patches,  or  '  milk  spots  ' 
(Ranvier)  in  it,  which  are  aggregations  of  corpuscles  like  lymph-corpuscles  ;  and  others,  which 
become  continuous  with  buds  given  off  from  existing  blood-vessels.  These  buds  ultimately 
become  channelled  to  form  blood-vessels. 

FRESH   BLOOD-VESSELS. 

Take  the  pia  mater,  washed  free  from  cerebral  matter,  and  tease  a  small  piece  on  a  slide  in 
salt  solution.  Cover.  An  examination  reveals  the  same  general  arrangement  as  described 
above,  but  the  nuclei  of  the  muscular  coat  of  the  arteries  requires  the  addition  of  acetic  acid 
to  bring  them  into  view.  Add  acetic  acid,  to  observe  this.  It  also  swells  up  the  adventitia, 
makes  it  clear  and  transparent,  and  the  elastic  lamina  when  present,  with  its  folds,  is  brought 
clearly  into  view — and  so  are  the  nuclei  in  the  capillary  wall.     Do  not  preserve  this. 

AORTA. 

PREPARATION. — Take  small  pieces  of  the  human  aorta  or  of  the  aorta  of  an  ox,  and 
place  it  in  picric  acid  for  twenty-four  hours.  Make  longitudinal  and  transverse  sections  in  the 
usual  way,  and  stain  them  with  picrocarmine,  taking  care  not  to  wash  the  sections  too  much  in 
water,  and  mount  them  in  Farrant's  solution. 

EXAMINATION  (L). — Observe  the  fusion  of  the  different  coats  and  the  large  amount  of 
elastic  fibres  and  plates  in  the  different  layers.  All  elastic  tissue  is  stained  yellow,  connective 
tissue  pink,  and  non-striped  muscle  light  reddish-brown,  so  that  these  different  tissues  are 
easily  distinguished  (H).  Note  the  numerous  layers  of  elastic  fibres  in  the  inner  coat ;  in  a 
transverse  section  they  appear  as  bright  yellow  granules  ;  in  the  longitudinal  as  yellow  lines. 
Note  the  large  amount  of  elastic  tissue  in  the  middle  coat,  where  it  occurs  in  the  form  of 
elastic  laminae. 

If  it  be  desired  to  see  the  lining  epithelium,  silver  (p.  xlv)  the  inner  surface  of  the  aorta  of 
an  animal  just  killed,  and  mount  a  thin  shaving  in  danuaar. 

ISOLATED    ELASTIC    LAMINA. 

PREPARATION. — By  far  the  best  method  is  the  one  I  described  several  years  ago  for  the 
isolation  of  elastic  tissue,  viz.,  to  digest  thin  strips  of  the  aorta  in  artificial  gastric  juice 
(p.  92)  till  everything  except  the  elastic  element  is  dissolved  (De  B.  Birch).  Wash  the  tissue,  to 
get  rid  of  acid,  and  mount  in  Farrant's  solution.  Examination  reveals  all  gradations,  from 
a  network  of  elastic  fibres  to  fenestrated  membranes  and  sheets  of  elastic  tissue.  The  elastic 
fibres  not  unfrequently  present  transverse  markings. 


X300 


Plate TXHe.aht  8^Blood^"essels 


Mirvfcarrx  Bi^os  .IxtK 


BLOOD-VESSELS.  51 


MIDDLE-SIZED   ARTERY. 

PREPARATION. — Place  a  moderate-sized  arterj' — say  the  femoral  or  posterior  tibial  of 
a  child,  or  the  basilar  artery  of  a  sheep  or  man — in  chromic  acid  and  spirit  for  a  week,  and 
make  transverse  sections.  Stain  one  section  with  logwood,  and  mount  it  in  dammar  ;  cover  ; 
another  with  picrocarmine,  and  mount  in  F~arrant's  solution. 

EXAMINATION  (L). — Obser\-e  the  inner  coat,  perhaps  with  the  layer  of  squames  lining  it. 
Observe  the  elastic  lamina,  as  a  clear,  yellow,  bright  line  thrown  into  folds  by  the  contraction 
of  the  middle  coat  which  surrounds  it.  If  a  large  artery  be  under  examination,  a  sub-epithelial 
layer  of  delicate  connective  tissue  may  be  found  between  the  epithelium  and  the  elastic 
lamina.  The  elastic  lamina  is  an  important  guide  in  disease  of  the  blood-vessels,  for  if 
changes  take  place  internal  to  it,  it  is  a  disease  of  the  inner  coat ;  if  immediately  external  to  it, 
the  disease  is  in  the  middle  coat.  The  student  should,  therefore,  familiarise  himself  with  its 
appearance,  on  account  of  its  important  pathological  relations.  The  middle  coat  consists 
chiefly  of  several  layers  of  circularly  disposed  non-striped  muscular  fibres.  The  nuclei  are 
long  ovals  and  of  a  violet  colour.  Between  the  fibres  may  be  found  elastic  tissue.  Outside 
this  note  the  adventitia,  consisting  mainly  of  white  fibrous  tissue  stained  blue,  and  containing 
elastic  fibres,  especially  in  its  inner  part.  {^Indicate  the  epitlwliiun,  elastic  lamina,  and  otiicr 
coats  in  PI.  IX.,  Fig.  8.) 

(H).  Study  the  elastic  lamina  ;  it  is  a  good  landmark  in  a  section.  On  its  inner  surface 
a  profile  view  may  be  obtained  of  the  lining  epithelium.     Observe  the  middle  and  outer  coat's. 

The  Epithelium  and  its  Cement-substance  in  blood-vessels  is  demonstrated  by  the  use  of  nitrate 
of  silver.      For  a  class  this  is  easily  managed,  thus  : — 

PREPARATION. — Kill  an  etherised  rabbit  by  bleeding.  Open  the  thorax  and  place  a 
cannula  in  the  aorta,  and  (after  making  an  opening  in  the  inferior  cava),  wash  out  the  blood- 
vessels with  distilled  water.  Then  inject  the  vessels  with  a  half  per  cent,  solution  of  silver  nitrate. 
Cut  out  the  intestines  and  wash  out  their  contents,  and  then  expose  them  and  their  mesentery 
in  a  mixture  of  equal  parts  of  spirit  and  water  to  sunlight  till  they  become  brown.  Select  either 
a  piece  of  the  mesentery  which  contains  blood-vessels,  or  what  answers  equally  well,  take  a  piece 
of  the  small  or  large  intestine  and  lay  it  on  a  slide  with  its  mucous  surface  upwards.  With  a 
scalpel  scrape  away  the  mucous  coat  and  mount  the  muscular  and  serous  coats  in  dammar. 

EXAMINATION  (L.  and  H). — Select  a  small  artery,  notice  the  silver  lines  running  trans- 
versely, indicating  the  disposition  of  the  circular  muscular  fibres,  and  inside  these  black 
silver  lines  mapping  out  narrow  lanceolate  areas  with  their  long  axis  in  the  axis  of  the  vessel 
which  are  the  squames  lining  the  artery.  Try  and  find  a  vein  and  compare  the  epithelium  of 
the  two  vessels.  Trace  an  artery  until  it  splits  up  into  capillaries,  and  study  the  silver  lines  in 
a  capillary.  These  indicate  that  it  is  made  up  of  elongated  flattened  endothelial  cells.  If  it 
be  desired  to  reveal  the  nuclei  in  these  plates,  a  section  should  be  stained  with  logwood. 
{^Indicate  the  silver  lines  in  a  capillary  in  PI.  IX.,  Fig.  4,  and  in  the  artery  in  Fig.  5.) 

VEINS. 

Exactly  the  same  methods  are  employed  for  bringing  into  view  their  structure.  Many 
sections  of  them  will  be  found  in  other  organs.  They  are  recognised  by  their  thin  walls,  the 
absence  of  so  perfectly  defined  an  elastic  lamina,  their  thinner  muscular  coat,  and  the  less 
distinct  demarcation  of  their  individual  coats. 

H  2 


52  PRACTICAL   HISTOLOGY. 


CIRCULATION  OF  THE  BLOOD. 

The  circulation  of  the  blood  may  be  studied  in  any  vascular  transparent  organ,  e.g.  web  of 
frog's  foot,  lung  of  frog,  and  also  in  the  mesentery.  For  the  student  it  is  best  to  study  the 
circulation  in  the  web  of  a  frog's  foot. 

METHOD. — Select  a  frog  with  a  slightly  pigmented  web,  and  with  a  hypodermic  .syringe 
inject  into  the  lymph-sac  under  the  skin  of  the  back  four  minims  of  a  quarter  per  cent,  watery 
solution  of  curara,  and  place  the  frog  under  a  bell  jar  till  its  motor  nerves  are  paralysed,  «>.  till 
it  ceases  to  move  when  it  is  pinched.  Place  the  frog  on  a  piece  of  card-board  one  and  a-half 
inches  wide  and  six  long,  with  a  triangular  slit  of  one-inch  base  cut  at  one  end  of  the  card.  Tie 
a  thread  round  the  tips  of  two  adjoining  toes,  e.g.  the  third  and  fourth.  Fix  the  threads  in  a 
slit  made  in  each  horn  of  the  cardboard,  and  stretch  the  web  gently  across  the  slit.  Moisten 
the  web,  place  the  card-board  on  the  stage  of  the  microscope,  and  fix  it  with  clips. 

EXAMINATION  (L). — Observe  the  arteries,  with  the  blood  moving  in  them  from  the  large  to 
the  smaller  vessels.  In  the  centre  of  the  stream  a  more  rapid  current,  and  between  it  and  the 
wall  of  the  vessel  the  slower  stream  or  lymph-space.  Select  a  vein,  known  by  its  thinner  walls 
and  the  direction  of  the  blood-stream,  and  notice  the  movement  from  the  smaller  to  the  larger 
vessels,  and  the  stream  slower  than  in  the  arteries.  Next  observe  the  network  of  capillaries, 
with  the  blood-corpuscles  moving  in  single  file.  Notice  also  the  pigment-cells,  some  of  them 
branched  ;  but  if  they  are  contracted  they  look  like  small  black  specks.  Cover  tlie  web  with  a 
fragment  of  a  cover-glass.  (H).  Study  a  capillary  ;  observe  its  thin  wall  and  the  passage  of 
the  corpuscles  in  single  file  through  it.  The  coloured  corpuscles  are  elastic,  as  shown  by  the 
way  in  which  they  twist  and  easily  become  distorted,  and  as  easily  regain  their  normal  shape. 
Select  a  small  vein  and  observe  the  colourless  corpuscles  dragging  lazily  along  in  the  lymph- 
space  and  adhering  to  the  wall  of  the  vessel. 

If  desired,  the  phenomena  of  inflammation  of  the  web  can  easily  be  studied  by  applying  a 
small  quantity  of  mustard  moistened  with  water,  or  some  otlier  irritant,  for  a  few  minutes. 
Wash  off  the  irritant  from  the  web  and  study  the  effects. 


53 


RESPIRATORY    ORGANS— LARYNX,    TRACHEA,    AND 

LUNGS. 

The  epiglottis  has  already  been  examined  (p.  i8).  The  preparation  may  be  referred  to 
again.  Its  anterior  surface  is  covered  by  stratified  epithehum,  and  so  is  the  posterior,  but  here 
the  submucous  coat  contains  adenoid  tissue,  and  a  large  number  of  mucous  glands  (p.  54), 
which  not  unfrequently  are  lodged  in  depressions  of  the  yellow  elastic  or  reticular  cartilage, 
which  constitutes  the  basis  of  the  organ.  Not  unfrequently  taste-bulbs  are  met  with  amongst 
the  epithelium  on  the  posterior  surface. 

The  arytenoid  cartilages  consist  of  yellow  fibro-cartilage,  and  sections  of  them  show  tran- 
sition stages  from  ordinary  hyaline  to  yellow  fibro-cartilage  (p.  19).  The  submucous  tissue 
contains  many  mucous  glands. 

If  desired  the  vocal  cords  may  be  hardened  in  chromic  acid  and  spirit  mixture,  and  vertical 
sections  made  in  the  ordinary  way. 


TRACHEA   AND   LUNGS, 

PEEPARATION. — Kill  an  etherised  cat  by  bleeding,  open  the  thorax,  and  allow  the  lungs 
to  collapse.  Tie  a  cannula  in  the  trachea,  and  with  a  syringe  distend  the  trachea  and  lungs 
with  a  mixture  of  chromic  acid  and  spirit  (p.  xxxi).  Suspend  the  lungs  in  a  large  quantity  of 
the  same  fluid.  Change  the  fluid  at  the  end  of  twenty-four  hours  ;  after  three  days  cut  the 
trachea  and  lungs  into  small  pieces,  and  place  them  in  fresh  hardening  fluid  for  a  week  or 
ten  days,  and  then,  after  washing  them  thoroughly,  transfer  them  to  alcohol  until  sections  are 
required.  Preserve  a  piece  of  human  trachea  in  the  same  way.  Distend  a  piece  of  human 
lung  and  preserve  it  as  above.  Make  transverse  and  longitudinal  sections  of  the  trachea, 
transverse  sections  through  a  bronchus  as  it  enters  the  lung,  and  sections  through  the 
pleura  and  subjacent  lung,  and  also  across  a  bronchus  within  the  lung-substance.  Preserve 
the  sections  in  preservative  fluid  (p.  xl)  until  they  are  required. 

Sections  of  such  an  organ  as  the  lung,  with  its  numerous  open  spaces,  ought  always  to  be 
made  after  steeping  in  gum,  which  fills  the  spaces  ;  and  after  freezing  the  organ  is,  as  it  were, 
solid,  and  the  gum  can  easily  be  dissolved  out  by  water.  Nothing  is  so  good  as  a  freezing 
microtome  for  making  sections  of  lung. 

The  lungs  are  covered  externally  by  a  serous  membrane,  ihc  pleura.  Its  epithelial  cover- 
ing can  easily  be  demonstrated  by  the  silver  process  (p.  xlv)  on  the  lungs  of  a  cat  just  killed. 
After  silvering,  a  superficial  slice  is  taken  and  mounted  in  dammar  or  glycerine.  If  the  lung 
be  silvered  while  it  is  distended  ad  maxijimm,  i.e.  in  a  state  of  inspiration,  the  cells  appear 
as  polygonal  squames  ;  if  it  be  collapsed,  i.e.  in  a  state  of  expiration,  the  cells  appear  slightly 
cubical  ;  showing  that  the  cells  are  soft  and  accommodate  themselves  during  life  to  the  changes 
of  size  in  the  air-vesicles  during  the  respiratory  acts. 


54  PRACTICAL    HISTOLOGY. 

TRANSVERSE  SECTION  OF  TRACHEA  OF  A  CAT. 
Stain  it  for  half  an  hour  in  picrocarmine  and  mount  in  Farrant's  solution. 
EXAMINATION  (L). — Observe  the  arc  of  hyaline  cartilage  forming  nearly  two-thirds  of  a 
circle  and  deficient  posteriorly.  The  cartilage  is  invested  by  a  fibrous  perichondrium  stained 
red.  It  is  continuous  with  the  layer  of  circularly  disposed  non-striped  muscular  fibres,  that 
connect  the  ends  of  the  cartilages  and  help  to  make  up  the  posterior  wall  of  the  trachea.  This 
muscle,  the  trachealis  muscle,  is  not  inserted  into  the  tips  of  the  cartilage-rings,  but  into  the 
outer  surface  of  the  fibrous  perichondrium,  some  distance  from  their  free  ends.  The  muscle  is 
easily  made  out,  for  it  has  a  dull  red  appearance.  Outside  the  cartilage  is  some  fat  and  con- 
nective tissue.  Observe  the  mucous  membrane  with  its  columnar  ciliated  epithelium,  and  under 
it  the  submucous  coat,  containing  many  mucous  glands,  most  of  the  acini  of  which  lie  internal 
to  the  cartilages,  and  it  may  be  some  adenoid  tissue.  The  sacs  of  each  mucous  gland  are  en- 
closed in  a  connective-tissue  capsule,  and  have  a  flattened  appearance  where  they  lie  inside  the 
cartilage,  but  are  round  or  oval  between  the  rings.  Between  the  different  groups  of  glands 
single  fat-cells,  or  small  groups  of  fat-cells,  may  be  seen.  {Indicate  the  general  chaj-acters  in 
FIX.,  Fig.  I.) 

(H).  Observe  the  epithelium  in  several  layers,  the  upper  layer  cylindrical  and  ciliated.     It 
rests  on  a  basement-membrane  not  easily  seen  in  the  trachea  of  the  cat,  though  easily  seen  in 
the  human  trachea,  where  it  plays  an  important  role  in  pathological  processes.     Amongst  the 
epithelium  are  goblet-cells  which  are  stained  of  a  deep  red.     Under  the  epithelium  observe 
beautiful   layers  of  elastic  fibres   arranged   longitudinally,    which   appear   as  rows   of  bright 
yellow  dots  lying  amongst  the  connective  tissue.     Observe  the  mucous  glands,  and  their  acini 
inside  the  cartilages,  and  trace  a  duct  to  its  opening  on  the  surface  in  a  funnel-shaped  manner. 
The  acini  of  the  glands  lie  in  the  submucous  coat,  which  in  addition  to  connective  tissue  con- 
tains  a  very  considerable  quantity  of  adenoid  tissue.     As  the  gland-ducts  pass  obliquely  to 
reach   the   surface,  it  is  difficult  to  get  a  complete  section  of  a  gland  with  its  excretory  duct. 
The  ciliated  epithelium  is  continued  a  short  distance  into  the  excretory  duct,  but  it  soon  gives 
place   to  moderately  tall  cylindrical  epithelium,  which  lines  the  duct.     The  alveoli  or  sacs  of 
the  gland  vary  in  appearance  according  to  whether  they  were  or  were  not  secreting  at  death.     If 
secretion  had  been  taking  place,  the  cells  lining  them   will  be  filled  with  a  clear  transparent 
mucus-like   mass,  and  few  granular  cells  are  to  be  seen.     If  there  has   been   no    secretion,  the 
gland-sacs  will  be  lined  by  a  regular  cubical  epithelium  of  a  more  or  less  granular  appearance. 
Observe   the  numerous   leucocytes   in   the  submucous  coat.     Observe  the  non-striped  muscle 
— trachealis— only  present  posteriorly  and  its  attachment  to  the  perichondrium  some  distance 
in  front  of  the  free  ends  of  the   tracheal  rings.     Outside  it   may  be  seen  transverse  sections 
of  nerves  and  nerve-ganglia.     As  the   mucous   membrane  is  very  vascular,  many  sections  of 
blood-vessels  will  be  seen  in  it.     The  cartilage  is  ordinary  hyaline  cartilage,  but  its  corpuscles 
present  a  characteristic  arrangement.     Under  the  perichondrium  the  cells  are  flattened,  and  lie 
parallel  to  the  surface  of  the  cartilage,  whilst  those  in  the  interior  are  elongated  or  oval  and  are 
placed  vertically  from  one  surface  to  the  other,  i.e.  across  the  long  axis  of  the  cartilage.     {Indi- 
cate the  epithelium,  glands,  and  part  of  the  cartilage  in  PI.  X.,  Fig.  2.) 

Double-stain  a  section  of  a  trachea  with  picrocarmine  and  iodine  green  (p.  xlv).    The  mucous 
glands,  adenoid  tissue,  and  cartilage  are  stained  green. 

Vertical  section  of  Trachea,  stained  with  picrocarmine  and  mounted  in  Farrant's  solution. 
EXAMINATION  (H).— Observe  the  elliptical  pieces  of  cartilage— the  sections   of  the  car- 
tilaginous rings  cut  vertically.     Observe  the  same   general  arrangement   as  described  in  the 


TRACHEA.  55 

transverse  section,  with  this  difference,  that  the  mucous  glands  are  seen  to  be  most  numerous 
between  the  cartilages,  and  the  longitudinally  disposed  elastic  fibres  under  the  epithelium 
can  now  be  detected.     Study  these  various  parts  under  a  high  power. 

HUMAN    TRACHEA. 

Transverse  section  of  a  Human  Trachea,  stained  with  picrocarmine  and  mounted  in  Farrant's 
solution. 

EXAMINATION  (L). — Observe  the  same  general  arrangement  of  cartilage,  glands,  tracheal 
muscle,  and  mucous  coat  as  described  in  the  cat's  trachea.  Notice,  however,  that  the  ciliated 
epithelium  is  usually  absent  at  some  parts,  from  the  difficulty  of  getting  perfectly  fresh  tis- 
sue to  preserve.  An  important  point  is  the  existence  of  a  well-marked  layer  of  tissue 
stained  of  a  deep  red — a  '  basement  membrane '  so  called.  It  lies  immediately  under  the 
epithelium  inside  the  longitudinally  disposed  layer  of  elastic  tissue.  It  is  a  marked  feature  in 
the  human  trachea,  and  plays  an  important  part  in  pathological  changes,  especially  in  those 
resulting  from  chronic  bronchitis.  Observe  the  mucous  glands  and  their  ducts  ;  the  acini  of 
many  of  them  lie  outside  the  trachealis  muscles,  so  that  their  ducts  have  to  perforate  it  to 
reach  the  surface  of  the  trachea.  The  largest  glands  lie  posteriorly,  and  they  present  the 
same  characters  as  previously  described,  their  appearance  varying  with  their  state  of  physio- 
logical activity. 

(H).  Where  the  superficial  epithelium  has  been  detached,  good  examples  of  developing 
epithelial  cells  may  be  studied.     Study  each  of  the  other  structures  in  detail. 

The  so-called  'basement  membrane,'  it  seems  to  me,  resembles  very  closely  a  similar  struc- 
ture which  exists  in  the  mucous  coat  of  the  cat's  stomach,  though  there  its  relation  to  the  gland- 
structures  is  different.  I  am  unable  to  discover  any  corresponding  structure  in  the  trachea  of 
the  rabbit,  cat,  or  dog.  Its  structure,  relations,  and  homologies  require  further  investigation. 
It  is  certainly  quite  different  from  the  basement-membrane  made  up  of  epithelial  cells,  and 
described  by  Dcbovc  as  occurring  under  the  epithelium  of  an  intestinal  villus,  the  bladder, 
trachea,  &c. 

Double-stain  a  section  of  human  trachea  with  picrocarmine  and  iodine  green. 

TRANSVERSE  SECTION  OF  A  BRONCHUS  AT  THE  ROOT  OF  THE  LUNG. 

Place  a  section  in  a  quarter  per  cent,  solution  of  osmic  acid  for  twenty-four  hours,  and 
mount  it  in  Farrant's  solution,  and  another  ought  to  be  stained  with  picrocarmine  and  simi- 
larly mounted. 

EXAMINATION  (L). — Observe  the  bronchus,  and  note  on  each  side  of  it  a  large  blood- 
vessel, the  one  a  branch  of  the  pulmonary  artery,  and  the  other,  on  the  opposite  side,  a  branch 
of  the  pulmonary  vein.  These  three  structures  lie  in  channels,  and  are  imbedded  in  connec- 
tive tissue,  and  are  thus  accurately  mapped  off  from  the  adjacent  vesicular  lung-tissue. 

Observe  the  bronchus.  Note  that  several  pieces  of  hyaline  cartilage  are  inserted  in  the 
bronchial  wall.  Outside  them  is  connective  tissue  often  containing  a  few  fat-cells — which  are 
blackened  in  the  osmic  acid  preparation.  Besides  this,  note  transverse  sections  of  small 
blood-vessels — the  bronchial  arteries,  and,  lying  near  them,  transverse  sections  of  nerves,  with 
perhaps  a  ganglion  in  their  course.  Observe  the  mucous  coat  thrown  into  folds,  and  under  it 
the  cut  ends  of  longitudinally  disposed  elastic  fibres.  Outside  this  a  continuous  ring  of  non- 
striped  muscular  fibre — the  bronchial  muscle.     It  is  perforated  here  and  there  by  the  ducts  of 


56  PRACTICAL   HISTOLOGY. 

the  mucous  glands,  whose  acini  lie  between  it  and  the  cartilages,  though  they  are  most  nume- 
rous where  no  cartilage  exists.  Perhaps  a  lymphatic  cord  may  be  found  in  the  submucous 
coat.  Observe  the  pulmonary  blood-vessels  and  compare  the  relative  thickness  of  their  coats, 
{Indicate  the  general  arrangement  in  PI.  X.,  Fig.  3.) 

(H).  The  epithelium  and  glands  have  exactly  the  same  structure  as  in  the  trachea.  The 
cut  ends  of  the  elastic  fibres,  stained  bright  yellow,  are  most  numerous  where  the  mucous  mem- 
brane is  raised  as  a  fold.  Study  the  fibrous  connective  tissue  mixed  with  elastic  fibres  which 
exists  outside  and  around  the  cartilages  and  blood-vessels.  Select  a  transverse  section  of  a 
nerve,  if  possible  with  a  ganglion  in  it.  The  ganglia  and  large  branches  of  nerves  will  be 
found  outside  the  cartilages,  and  usually  accompanied  by  one  or  more  small  blood-vessels 
(bronchial).  Observe  the  ganglionic  cells,  large  oval  corpuscles  with  a  distinct  nucleus  and 
nucleolus,  and  between  or  around  them  the  cut  ends  of  nerve-fibres  medullated  and  non- 
medullated,  the  whole  surrounded  by  a  fibrous  sheath.  {Indicate  a  small  part  of  the  loall  of  a 
bronchus  in  PI.  XI.,  Fig.  i.) 

Vertical  Section  through  the  Pleura  and  the  Subjacent  Vesicular  Tissue.— Stain  with  logwood 
and  mount  in  dammar. 

EXAMINATION  (L). — Observe  the  pleura,  made  up  of  fibrous  tissue  stained  blue,  and  note 
its  normal  thickness.  From  its  under  surface  it  sends  into  the  lung,  at  regular  intervals,  fine 
processes  of  connective  tissue— the  interlobular  septa — i.e.  processes  between  adjoining  lobules. 
It  may  be  possible  to  trace  the  connection  of  these  septa  with  the  connective  tissue  around  a 
bronchus.  They  contain  many  lymphatics,  and  are  often  pigmented  from  the  presence  of 
charcoal  or  soot,  especially  in  the  human  lung.  They  are  of  great  importance  pathologically, 
and  this  connection  of  the  pleural  connective  tissue  with  that  which  enters  the  lung  at  its  root 
is  also  important,  e.g.  in  the  production  of  dilatation  of  the  bronchi  and  other  pathological 
processes.  Here,  as  in  other  organs  it  is  important  to  bear  in  mind  the  connection  of  the  con- 
nective tissue  of  the  capsule  with  that  in  the  interior  of  the  organ.  Observe  now  the  air- 
vesicles,  cut  in  every  direction,  but  notice  that  those  placed  most  superficially,  i.e.  next  the 
pleura,  are  somewhat  pyramidal  or  conical  in  shape,  and  have  their  bases  directed  towards  the 
pleura  ;  the  air-vesicles  elsewhere  are  polygonal  in  shape,  and  may  or  may  not  have  a  base  to 
them,  which  depends  on  the  line  of  section.  They  are  separated  from  each  other  by  a  very 
small  quantity  of  connective  tissue,  and  by  elastic  fibres,  which  form  a  network  round  each 
air- vesicle.  Here  and  there  a  transverse  section  of  a  small  bronchus  may  be  found.  It  is 
recognised  by  the  cubical  epithelium  lining  it,  and  by  the  presence  of  cartilage  or  glands  in 
its  walls.  In  the  vesicular  structure  search  for  a  transverse  section  of  an  infundibulum.  The 
nuclei  seen  in  the  air-vesicles  belong  to  the  squames  lining  them,  and  those  forming  the 
capillaries  in  their  walls.  Search  for  a  small  bronchus  opened  into  longitudinally,  and  observe 
its  dichotomous  division  and  its  expansion  into  air- vesicles.  {Indicate  the  pleura,  and  the  shape 
of  the  air-vesicles,  in  PI.  XI.,  Fig.  2.) 

(H).  Observe  thepleu7-a,  which  consists  of  two  layers  ;  the  superficial  one  contains  a  con- 
siderable number  of  elastic  fibres,  and  the  lower,  or  sub-pleura,  consists  of  looser  connective 
tissue,  which  is  continued  into  the  lung  between  the  lobules  as  interlobular  septa.  In  it  may 
be  seen  sections  of  the  pleural  blood-vessels  and  lymphatics,  the  latter  appearing  as  fine  slits. 
The  endothelium  on  the  surface  of  the  pleura  is  not  visible. 

Observe  an  air-vesicle.  Note  its  outline— rounded  or  polyhedral.  If  its  thin  edge  be  seen 
in  profile,  observe  the  squamous  epithelium  lining  it,  though  it  is  better  seen  where  a  portion 
of  the  wall  of  an  alveolus  is  flat  in  the  field.     In  the  alveolar  wall  trace  the  branching  elastic 


Plate  Z.Trachfa  ^Ijjtig 


lantern,  Bros  .]itK 


LUNGS.  57 

fibres,  and  carefully  distinguish  the  plexus  of  capillaries,  sometimes  containing  a  few  blood-cor- 
puscles. {Indicate  the  epithelium  and  elastic  fibres  in  PI.  XL,  Fig  3.)  In  the  capillary  wall 
observe  the  nuclei,  which  are  not  to  be  confounded  with  those  of  the  squames  lining  the 
air-vesicle.  In  the  cat's  lung,  fusiform  nuclei,  indicating  the  existence  of  non-striped  muscle 
between  the  vesicles,  are  to  be  detected.  The  amount  of  non-striped  muscle  between  the  air- 
vesicles  is  sometimes  greatly  increased  in  amount  in  certain  diseases.  It  is  easy  to  see  this 
in  the  lungs  of  a  cat  suffering  from  the  presence  of  the  ova  of  a  worm  {Olhdaniis  triciispis)  in 
the  lungs  (Stirling).  Select  a  very  small  broncluis  cut  longitudinally.  Both  cartilage  and 
glands  have  disappeared  from  its  walls,  and  the  epithelium  lining  it  is  not  so  tall  as  in  larger 
bronchi.  Trace  it  into  an  alveolar  duct,  which  passes  into  the  infundibulum,  into  which  the 
terminal  alveoli  open  on  all  sides.  In  the  alveolar  duct  and  infundibulum  the  epithelium  con- 
sists of  low  polyhedral  cells  without  cilia.  The  walls  of  the  alveolar  ducts  and  infundibula 
contain  many  non-striped  muscular  fibres,  disposed  circularly,  which  are  in  direct  continuity 
with  the  bronchial  muscle. 

Pierocarmine  Preparation. — Stain  a  similar  section  with  picrocarmine,  and  mount  it  in 
Farrant's  solution.  After  several  days  it  shows  the  above  details,  even  better  than  the  dammar 
preparation, 

Osmic  Acid  Preparation. — Place  a  section  of  lung,  hardened,  as  above,  in  chromic  acid  and 
spirit  mixture,  in  a  quarter  per  cent,  solution  of  osmic  acid  for  twenty-four  hours,  and,  after 
washing,  mount  it  in  Farrant's  solution.  It  shows  the  above  details  most  beautifully  ;  the 
elastic  fibres  stand  out  very  prominently,  and  their  arrangement  is  easily  made  out. 

Vertical  Section  of  Human  Pleura  and  Lung. — Prepared  as  above,  and  similarly  mounted. 
Note  the  more  conspicuous  interlobular  septa,  often  pigmented  from  the  deposition  of  charcoal 
particles  in  the  lymphatics.  Compare  other  parts  of  the  section,  and  note  especially  the 
branching  character  of  the  elastic  tissue,  for  the  physician  is  sometimes  called  on  to  recognise 
these  fibres  in  the  sputum  of  a  person  suffering  from  gangrene  of  the  lung. 

HUMAN    FOITAL   LUNG. 

Prepare  the  lung  of  a  foetus — preferably  one  that  has  not  respired — in  the  same  way,  with 
chromic  acid  and  spirit  mixture,  as  directed  for  adult  lung.  In  ten  days  it  will  be  ready 
for  cutting.  Or,  use  picric  acid  solution  as  the  injecting  and  hardening  medium, which  will 
harden  it  in  two  days.  Make  sections  across  a  lung,  and,  after  staining  with  picrocarmine, 
mount  in  Farrant's  solution. 

EXAMINATION  (L). — Observe  the  distinct  lung-lobules,  each  covered  on  its  free  broad  end 
by  a  relatively  thick  pleura,  and  separated  from  its  neighbouring  lobule  by  thick  interlobular 
septa,  whose  continuity  with  the  adventitia  or  connective  tissue  of  the  intra-pulmonary  bronchus 
can  easily  be  made  out.  In  each  interlobular  septum  note  the  large  open  spaces,  which  are 
sections  of  the  interlobular  lymphatics.  Study  the  small  non-distended  air-vesicles,  and  note 
(H)  that  they  are  lined  by  low  cubical  cells,  not  yet  converted  into  squames  by  the  distension 
of  the  lung  as  the  result  of  the  act  of  inspiration.  {Indicate  the  pleura  and  interlobular  septa 
in  PI.  XL,  Fig.  6.) 

SQUAMOUS    EPITHELIUM    OF    THE   AIR-VESICLES. 

PREPARATION.— Remove  the  lungs  of  a  cat  or  kitten  from  the  thorax,  and  distend  them 
with  a  quarter  per  cent,  solution  of  nitrate  of  silver,  and  keep  them  distended  by  tying  a 
string  round  the  trachea.    Place  them,  with  a  weight  attached,  in  alcohol  until  required.    When 

I 


58  PRACTICAL   HISTOLOGY. 

requircc!,  cut  the  lunt^  into  pieces,  and  make  sections  by  freezing.  Expose  the  sections  to  the 
action  of  light,  until  they  become  brownish  in  colour.  Stain  a  section  with  picrocarmine  for 
twenty-four  hours,  and  mount  it  in  glycerine  ;  and  another  with  logwood  for  a  few  minutes, 
and  mount  it  in  dammar. 

EXAMINATION  (L). — Observe  the  same  general  arrangement  of  the  air-vesicles  as  already 
described.     Select  an  air-vesicle  where  a  portion  of  its  wall  is  seen  on  the  flat,  and  examine. 

(H).  Observe  the  silver  lines,  indicating  the  existence  of  epithelium  (squamous),  lining  the 
vesicle.  These  lines  bound  polygonal  areas.  Between  these  larger  and  clear  areas  notice 
small  brown  granular  areas  or  polyhedral  cells,  in  groups  of  two  or  three,  some  of  them 
with  a  nucleus  stained  red  or  blue.  These  are  young  epithelial  cells,  and  are  thicker  than 
the  squames  or  placoids.  Transition  forms  exist  between  them  and  the  clear  squames. 
During  a  maximum  distension  of  the  lung  they  yield  and  become  flattened  out.  In  a  well- 
distended  alveolus,  narrow  angular  apertures,  especially  in  the  cement  lines,  are  to  be  seen. 
These  are  the  so-called  pseudostomata,  or  small  apertures  which  lead  into  the  lymph-canalicular 
system  of  the  alveolar  wall.     {^Indicate  the  squames  and  small  granular  cells  in  PI.  XL,  Fig.  4.) 

BLOOD-VES.SELS    OF    THE    LUNGS. 

The  branches  of  the  pulmonary  artery  break  up  into  capillaries,  which  are  distributed  on 
the  walls  of  the  air-vesicles.  At  the  root  of  the  lung  the  branches  of  the  pulmonary  artery 
and  vein  and  bronchus  (p.  55)  are  found  together, — the  bronchus  in  the  middle,  with  a  blood- 
vessel on  each  side  of  it, —  within  the  lung  the  pulmonary  vein  pursues  a  separate  course. 
The  bronchial  arteries  enter  the  lung  at  its  root,  and  run  in  the  adventitia  of  the  large  bronchi, 
which  they  supply  with  blood.  Sections  of  them  will  be  found  near  the  nerve-trunks  at  the 
root  of  the  lung  (p.  55). 

PREPARATION. — Make  sections  of  a  lung  whose  blood-vessels — pulmonary  artery,  or 
vein,  or  both — have  been  injected  with  a  carmine  gelatine  (p.  li)  or  Prussian  blue  (p.  li)  mass, 
and  mount  them  in  dammar. 

EXAMINATION  (L). — Notice  the  extremely  dense  plexus  of  capillaries  on  the  walls  of 
each  air-vesicle.  Trace  a  small  arteriole  from  a  large  branch  of  the  pulmonary  artery,  and 
note  that  it  may  supply  two  or  three  adjacent  alveoli  ;  while  the  efferent  vein  usually  passes 
off  at  the  other  side.     (^Indicate  the  arrangement  of  the  capillaries  in  PI.  XL,  Fig.  5.) 

(H).  Study  the  capillaries  ;  note  the  dense  network  of  more  or  less  wavy  capillaries,  and 
especially  on  a  septum  between  two  adjacent  alveoli.  A  twisted  capillary  may  be  seen  lying 
at  one  time  in  one  alveolus,  at  another  in  the  adjoining  alveoli.  The  space  between  the 
capillaries  varies  according  as  the  lung  has  been  kept  distended  or  not.  In  preparations  which 
have  been  placed  in  alcohol,  the  air-vesicles  shrink  considerably,  and  hence  the  alveolar 
capillaries  appear  relatively  close  to  each  other.  The  large  arterial  and  venous  branches  lie 
in  the  interlobular  septa,  which  are  continuous  with  the  adventitia. 

LYMPHATICS    OF   THE    LUNG. 

These  are  very  numerous,  and  form  three  systems  : — (i)  the  subpleural  lymphatics  occur 
in  the  deep  layer  of  the  pleura,  and  communicate  with  the  pleural  cavity  on  the  one  hand, 
and  have  a  direct  connection  with  the  lymphatic  canalicular  system,  which  lies  in  the  alveolar 
wall,  and  which  communicates  with  the  pseudo-stomata  of  the  air-vesicles.  Branches  of 
this   system    pass   through  the  interlobular  septa    to   reach   (2)  the  perivascular  lymphatics 


Plate  XI.  LunCt 


iviiitem  Bros.iith 


LUNGS.  59 

which  accompany  the  branches  of  the  pulmonary  artery  and  vein  ;  (3)  the  peribronchial 
lymphatics  which  occur  in  the  adventitia  of  the  bronchi,  communicate  freely  with  the  peri- 
vascular lymphatics,  and  run  towards  the  bronchial  glands.  In  the  walls  of  the  bronchi  of 
the  cat  large  masses  of  adenoid  tissue  or  lymph-cords  are  easily  found  (Klein,  Sanderson,  and 
Stirling).     They  are  not  nearly  so  abundant  in  the  human  lung. 

PEEPARATION. — The  lymphatics  of  the  bronchi  may  be  injected  by  the  puncture  method  ; 
but  a  good  idea  of  their  arrangement  may  be  obtained  by  the  student  from  the  lungs  of  a 
coal-miner  who  has  suffered  from  anthracosis.  The  particles  of  soot  and  charcoal  are 
carried  into  the  lymphatics,  which  therefore  appear  black.  Make  a  vertical  section  through 
the  pleura  and  lung-tissue  of  such  a  lung,  which  has  been  hardened  in  alcohol,  and  mount  it 
in  Farrant's  solution. 

EXAMINATION  (L).— Observe  the  black  pigment  distributed  in  the  lung.  Note  that  the 
superficial  layer  of  the  pleura  has  no  pigment  in  it,  though  the  deep  layer  is  markedly  pig- 
mented. Valves  prevent  the  passage  of  the  pigment  from  the  deep  into  the  superficial 
lymphatics  of  the  pleura.  Trace  from  the  pleura  inwards  strands  of  pigment  in  the  inter- 
lobular septa,  in  the  outer  coats  of  the  arteries  and  bronchi,  and  it  may  be  seen  here  and  there 
between  the  air-vesicles,  and  also  in  the  adventitia  of  the  bronchi  and  arteries.  (Compare 
Klein's  'Anatomy  of  the  Lymphatic  System,'  pt.  ii.,  1875.) 

The  Nerves  of  the  Lung  are  very  numerous,  and  enter  it  with  the  bronchi.  As  already 
indicated,  they  may  be  seen  in  a  section  of  the  root  of  the  lung.  They  lie  outside  the 
cartilages  in  the  bronchial  adventitia,  and  are  accompanied  by  branches  of  the  bronchial 
arteries.  Five  or  six  strands  of  nerve-fibres  may  be  found  in  one  section.  Many  ganglia 
(Remak,  Klein,  and  Stirling)  are  found  in  their  course,  and  are  often  seen  in  section.  They 
can  be  easily  isolated  from  a  bronchus  with  the  aid  of  a  dissecting  microscope  (p.  xxxv). 

The  Nerves  of  the  Trachea. — I  have  recently  discovered  a  large  number  of  ganglia  inter- 
calated in  the  course  of  the  nerves,  lying  just  outside  the  posterior  muscular  wall  of  the  trachea. 
Some  of  them  may  be  seen  in  sections  of  the  trachea. 


13 


6o  PRACTICAL    HISTOLOGY. 


ALIMENTARY   CANAL,    AND    THE    GLANDS 
CONNECTED    WITH  IT. 

THE    LIPS. 

PREPARATION. — Place  small  pieces  of  the  lips  of  a  dog  or  cat  in  chromic  acid  and  spirit 
mixture  for  two  weeks,  and,  when  they  are  hardened,  make  vertical  sections  of  them  in  the 
ordinary  way.     Stain  a  section  with  picrocarmine,  and  mount  it  in  Farrant's  solution. 

EXAMINATION  (L).— Observe  the  hair-follicles  on  the  outer  surface  of  the  lip,  and  the 
inner  surface  covered  by  stratified  squamous  epithelium,  and  if  the  animal's  skin  was  pigmented 
the  deeper  layers  of  the  epithelium  contain  black  pigment  or  melanin.  Between  these  two 
surfaces  observe  the  transverse  sections  of  the  orbicularis  oris  muscle,  its  muscular  bundles 
surrounded  by  connective  tissue,  continuous  with  the  connective  tissue  of  the  rest  of  the  lip. 
All  the  connective  tissue  is  red,  the  glands  and  muscle  yellowish,  and  their  nuclei  deep  red. 
Numerous  sections  of  blood-vessels  and  nerves  are  seen.  (H).  Place  the  stratified  epithelium 
in  the  field  of  the  microscope  and  examine  it.  Observe  the  epithelium,  and  study  particularly 
the  prickle-cells  and  the  pigment  in  the  cells  of  the  deeper  layers.  The  various  layers  of  a 
hair-follicle  are  easily  made  out,  and  the  sebaceous  glands  opening  into  them  are  easily 
studied  (compare  hair-follicles,  p.  93). 

TONGUE. 

The  muscular  substance  of  the  tongue  is  enclosed  in  a  mucous  membrane,  covered  with 
stratified  epithelium.  The  mucous  membrane  on  the  back  and  sides  of  the  tongue  is  provided 
with  papillae — filiform,  fungiform,  and  circumvalate — and  on  these  are  small  secondary  papillae. 
There  is  a  thin  sub-mucous  layer,  whose  connective  tissue  is  continuous  with  that  surrounding 
the  muscular  substance.  The  muscular  substance  consists  of  longitudinal,  transverse,  and 
vertical  bundles  of  striated  muscle.  In  the  posterior  part  of  the  dorsum  of  the  tongue  are  to 
be  found  mucous  and  serous  glands  and  adenoid  tissue,  and  on  the  sides  of  the  tongue  the 
papillae  foliatse,  in  which  lie  the  taste-bulbs. 

PREPARATION. — Harden  the  tongue  of  a  cat,and  also  a  part  of  the  human  tongue,  in  chromic 
acid  and  spirit  mixture  for  two  weeks,  and  make  transverse  sections.  Stain  one  with  logwood 
and  mount  it  in  dammar,  and  stain  another  with  picrocarmine  and  mount  it  in  Farrant's  solution. 

EXAMINATION  (L). — Observe  the  papillae,  confined  to  the  dorsum  of  the  tongue,  beset 
with  secondary  papillae,  and  covered  with  epithelium  ;  beneath  these  the  scanty  sub-mucous 
coat,  containing  sections  of  blood-vessels.  In  the  middle  of  the  muscular  substance  a  verti- 
cally placed  septum,  and  muscular  fibres  running  transversely  out  from  it.     Observe  muscular 


TONGUE.  6 1 

fibres  disposed  vertically,  and  terminating  above  in  tendons  continuous  with  the  connective 
tissue  of  the  mucous  membrane.  Immediately  under  the  mucous  membrane  of  the  dorsum 
notice  the  transverse  sections  of  longitudinal  muscular  fibres.  Between  the  muscular  fibres  many 
fat-cells  are  seen,  and  excellent  sections  of  nerves,  and  the  branches  of  the  ranine  artery  are 
found  in  the  lower  part.  {Indicate  the  general  arrangement  of  the  part^  in  one  half  of  V\.  XII., 
Fig.  I.)  (H).  Study  the  papilla;,  and  the  epithelium  covering  them.  The  upper  layers  of 
the  epithelium  are  often  horny,  especially  in  animals.  Notice  the  transverse  sections  of 
muscular  fibres — polygonal  in  shape — with  the  nuclei  placed  just  under  the  sarcolemma  (PI. 
XII.,  Fig.  2). 

POSTERIOR  PART  OF  THE  TONGXIE.—Prepare  this  just  as  the  anterior  part,  and 
make  vertical  sections.  Here  the  mucous  membrane  is  much  thicker  and  looser,  and  contains 
many  secreting  glands,  lymphatic  follicles,  and  diffuse  adenoid  tissue.  Stain  a  section  with 
logwood  and  mount  it  in  dammar. 

EXAMINATION  (L). —  Perhaps  the  section  may  have  passed  through  a  circumvalate 
papilla..  Study  specially  the  secretifig  glands,  which  are  of  two  kinds,  mucous  and  scro7is 
(v.  Ebner).  The  mucous  (L  and  H)  glands  are  like  similar  glands  in  the  mouth  and  oesopha- 
gus (p.  65).  Each  gland  consists  of  a  duct  and  a  secreting  part.  The  duct  passes  vertically 
and  opens  on  the  free  surface,  often  with  a  funnel-shaped  mouth.  It  has  a  wide  lumen,  and  a 
membrana  propria  with  oval  nuclei,  lined  by  a  layer  of  columnar  epithelium.  The  duct 
branches  and  terminates  in  the  secretory  part,  which  consists  of  more  or  less  convoluted  tubes, 
so  that  each  gland  is  a  compound  tubular  gland.  The  cells  lining  the  secretory  alveoli  consist 
of  a  single  layer  of  low  columnar  epithelium,  and  their  appearance  is  very  characteristic, 
especially  when  they  are  distended.  They  are  clear  and  transparent,  with  a  thin  oval  nucleus 
pushed  to  one  side.  They  contain  a  delicate  network  of  fibrils  distended  with  '  mucigen,' 
which  yields  mucin.  During  a  state  of  exhaustion  these  cells  are  more  '  granular.'  The 
serous  glands  (L  and  H)  always  bear  a  relation  to  the  taste-bulbs,  and  these  ducts  open  into 
grooves  which  contain  these  bulbs.  The  secreting  epithelium  is  columnar  with  a  spherical 
nucleus,  and  the  protoplasm  is  very  granular,  due  to  the  existence  of  a  dense  fibrillar  network. 
This  markedly  granular  character  at  once  distinguishes  them  from  the  clear  transparent  cells 
of  the  mucous  glands.  They  seem  to  secrete  a  watery  fluid.  In  the  same  section  patches  of 
adenoid  tissue  may  be  found. 

Double-Staining  of  the  Tongue  with  Logwood  and  Iodine  Green. — Stain  a  vertical  section  of 
the  posterior  part  of  the  tongue  lightly  in  logwood,  wash  it  thoroughly,  and  then  stain  it 
slightly  in  a  weak  watery  solution  of  iodine  green  (p.  xlv)  ;  mount  in  dammar.  The  striking 
feature  is  that  the  acini  of  the  mucous  glands  are  stained  of  a  bright  green,  while  the 
epithelium  in  the  efferent  duct  is  of  a  logwood  tint.  A  high  power  reveals  the  nuclei  stained 
with  logwood.  The  serous  glands  do  not  take  on  any  of  the  green  pigment,  so  that  the  two 
kinds  of  glands  stand  out  in  striking  contrast.  Make  a  similar  preparation  with  picrocarmine 
and  iodine  green  as  indicated  for  the  small  intestine  (p.  69)  ;  mount  in  dammar.  All  the  muscle 
will  be  yellowish-red,  the  connective  tissue  deep  red,  and  the  adenoid  tissue  and  mucous  glands 
bright  green. 

Blood-vessels  of  the  Tongue. — Mount  a  vertical  section  of  an  injected  tongue  in  dammar.  The 
blood-vessels  present  nothing  peculiar.  Trace  branches  into  the  papill<-E.  If  the  papilla  be  simple 
it  contains  a  single  capillary  loop  ;  if  compound,  each  secondary  papilla  contains  a  similar  loop. 
{Fill  in  the  blood-vessels  in  one  half  of  Y\.  XII.,  Fig.  i.) 

Nerves  of  the  Tongue. — Many  sections  of  the  hypoglossal  nerve  will  be  found  near  the 


62  PRACTICAL   HISTOLOGY. 

ranine  artery  in  any  of  the  above  sections.  In  the  course  of  the  Hngual  and  glosso-pharyngeal 
nerves,  not  unfrequently  fine  ganglia  are  seen.  I  have  often  seen  them  in  sections,  and 
isolated  them  from  these  nerves  (Remak,  Kolliker,  Stirling). 

TASTE-BULBS.— The  papilla;  foliatse  on  the  sides  of  the  back  part  of  the  rabbit's  tongue 
are  hardened  in  the  same  way  as  the  tongue.  Make  vertical  sections  through  the  leaflet-like 
papilhe.  After  staining  them  with  logwood,  mount  in  dammar.  They  are  also  found  on  the 
circumvalate  papilla;,  but  the  papillee  foliatae  are  preferable.  They  are  also  found  on  the  pos- 
terior surface  of  the  epiglottis. 

EXAMINATION  (L). — Observe  the  flattened  papillae  lying  side  by  side  ;  and  on  each  side 
of  a  papilla  the  four  oval  or  flasklike  taste-bulbs.  At  the  base  of  the  papillaj  the  ducts  of 
serous  glands  (p.  6i)  may  be  met  with  (PL  XII.,  Fig.  3). 

(H).  Study  a  taste-bulb  made  up  of  narrow  epithelial  cells  arranged  like  the  staves  of  a 
barrel,  investing  modified  epithelial  cells,  each  provided  with  a  fine  process,  which  projects 
through  an  opening  at  the  upper  part  of  the  investing  cells  (PI.  XII.,  Fig.  4). 

The  'inner'  and  'outer'  cells  may  easily  be  isolated  by  teasing  out  a  small  piece  of  a 
papilla,  hardened  in  a  quarter  per  cent,  osmic  acid  solution. 


TOOTH. 

UNSOFTENED    TOOTH. 

PREPARATION. — The  same  as  that  employed  for  dense,  dry  bone  (p.  n).  It  is  better 
to  purchase  a  section  ready-made  than  to  spend  time  in  grinding  down  a  section. 

EXAMINATION  of  a  vertical  section  (L). — Observe  the  dentine  or  ivory  forming  the  great 
mass  of  the  tooth  and  surrounding  the  pulp-cavity  in  the  centre,  the  enamel  covering  the 
crown  and  sides  of  the  dentine  and  the  crusta  petrosa  or  cement,  a  layer  of  bone  without 
Haversian  canals  covering  the  fang.  In  the  enamel  are  a  number  of  concentric  dark  lines— 
'  contour  lines  ' — running  across  it.  In  the  dentine,  note  the  wavy  lines  and  their  varying 
direction — the  dentinal  tubules.  They  stand  out  clearly  because  they  are  filled  with  air. 
Arched  incremental  lines  may  be  seen  running  across  the  dentine. 

(H).  Dentine. — In  the  homogeneous  matrix  the  dentinal  tubules,  which  open  by  one  end 
into  the  pulp-cavity  and  run  out  with  a  wavy  course  through  the  dentine,  dividing  dichoto- 
mously,  and  giving  off  many  anastomosing  lateral  branches,  end  in  the  outer  part  of  the 
dentine,  either  in  loops,  or  they  open  into  irregular  spaces — the  interglobular  spaces. 

Cement  or  Crnsta  Petrosa. — Like  bone  but  without  Haversian  canals.  Some  of  the  lacunae 
may  open  into  the  interglobular  spaces. 

Enamel. — Observe  the  transversely  striated  polyhedral  prisms  set  in  groups  on  the  dentine. 
If  cut  transversely,  they  are  seen  to  be  hexagonal.  Not  unfrequently  cracks  are  seen  in  the 
enamel  where  it  rests  on  the  dentine,  and  these  cracks  appear  black  from  being  filled  wjth 
air. 

SOFTENED   TOOTH. 

PREPARATION. — Take  the  lower  jaw  of  a  cat  or  dog,  and  carefully  free  it  from  its  muscles, 
but  retain  the  periosteum.     Cut  it  in  pieces  of  an  inch  long  with  a  saw,  and  place  the  pieces  in 


Plate  M  Tongue  &  Tooth 


>Entern.  Bi-os  EtH. 


TOOTH.  63 

a  large  amount  of  chromic  acid  and  nitric  acid  fluid  until  they  are  sufficiently  decalcified,  which 
may  be  ascertained  by  piercing  them  with  a  needle.  Decalcification  takes  place  in  two  to  three 
weeks  if  the  fluid  be  frequently  renewed.  Wash  the  pieces  frequently  in  water  to  remove  the 
surplus  acid,  and  then  transfer  them  to  spirit  until  sections  are  required.  Make  a  number 
of  vertical  sections  through  the  jaw,  and  the  tooth  implanted  therein.  Make  a  number 
of  sections  across  the  long  axis  of  a  tooth  and  keep  them  till  they  are  required.  In  the 
process  of  softening,  the  enamel  disappears,  owing  to  the  small  amount  of  organic  matter 
present.  Stain  a  vertical  section  and  a  transverse  section  in  picrocarmine,  and  mount  each 
in  Farrant's  solution. 

Vertical  section  of  a  softened  tooth.  EXAMINATION  (L). — Observe  the  absence  of  enamel, 
though  the  general  distribution  of  the  parts  of  the  tooth  is  the  same  as  in  the  unsoftened 
tooth.  Notice  the  tooth  planted  in  the  alveolus,  which  is  lined  by  a  membrane  stained  red — 
t\\Q  periodontal  vianbrajie — which  serves  as  a  periosteum  for  the  bony  wall  of  the  alveolus. 
Trace  its  continuity  with  the  fibrous  tissue  of  the  mucous  membrane  of  the  gum.  In  the  lower 
jaw-bone,  observe  the  medullary  cavity  with  branches  of  a  nerve  and  blood-vessels.  They  are 
very  apt  to  fall  out,  however.  The  pulp-cavity  is  seen  to  contain  the  pulp.  {Indicate  the  general 
arrangement  of  the  parts  in  PI.  XII.,  Fig.  5.) 

(H).  Observe  the  dentinal  tubules,  which  are,  however,  less  distinct  than  in  the  dry  tooth  ; 
the  matrix  is  stained  yellow.  Here  and  there  transverse  sections  of  the  tubules  may  be  seen  ; 
they  appear  as  fine  dots  in  a  homogeneous  matrix.  Notice  the  crusta  petrosa  with  its 
bone-corpuscles.  In  the  pulp-cavity  traces  of  blood-vessels,  delicate  connective  tissue,  and, 
it  may  be,  a  layer  of  columnar  cells — the  odontoblasts — may  be  found  lying  on  the  dentine. 
Place  a  similar  section  in  one  per  cent,  osmic  acid  for  four  hours,  mount  it  in  Farrant's 
solution,  and  examine.     It  shows  the  same  details  more  clearly. 

Transverse  section  of  a  softened  tooth.  EXAMINATION  (L  and  H). — Observe  the  pulp- 
cavity  and  the  dentinal  tubules  radiating  from  it,  and  many  of  them  cut  transversely. 

DEVELOPING   TOOTH. 

PEEPARATION.  —  Place  the  head  of  a  new-born  rat,  kitten,  or  puppy  in  a  large  quantity 
of  Mtiller's  fluid  for  two  days,  and  then  into  a  sixth  per  cent,  solution  of  chromic  acid  for  two 
days.  At  the  end  of  another  two  days,  substitute  for  it  the  chromic  and  nitric  acid  mixture 
(p.  xxxiii)  until  complete  decalcification  takes  place.  Make  vertical  sections  through  the  jaw, 
and  pass  them  through  a  five  per  cent,  solution  of  sodic  bicarbonate,  to  remove  all  trace  of  acid. 
Stain  a  section  with  picrocarmine,  another  must  be  left  in  one  per  cent,  osmic  acid  for  several 
hours,  and  mount  both  in  Farrant's  solution. 

EXAMINATION  (L). — Observe  the  general  shape  of  a  tooth-germ  still  imbedded  in  the 
gum.  Note  the  dentine  and,  it  may  be,  the  cap  of  enamel  ;  inside  the  dentine,  the  large 
mass  of  tissue  which  becomes  the  pulp. 

(H).  Lining  the  dentine,  observe  a  layer  of  columnar  cells — the  odontoblasts— which  give 
off  fibres — the  fibres  of  Tomes — which  pass  into  the  dentinal  tubules. 


64  PRACTICAL   HISTOLOGY. 


SALIVARY    GLANDS. 

The  salivary  glands  do  not  all  possess  the  same  structure,  nor  do  corresponding  glands  in 
different  animals  exactly  resemble  each  other  ;  hence  the  necessity  for  examining  the  various 
glands  of  different  animals.  There  are  three  kinds  of  salivary  glands,  which  differ  in  structure 
and  in  the  nature  of  their  secretion  : — 

(a)  True  Salivary  Glands — as  the  sub-maxillary  gland  of  rabbits  and  the  parotid  of  man 
and  some  mammals. 

{b)  True  Mucous  Glands — as  the  sub-maxillary  glands  of  cat  and  dog,  and  the  human 
sublingual. 

(c)  Muco-salivary  Glands — as  the  sub-maxillary  gland  of  man  and  guinea-pig. 

They  are  all  compound  tubular  glands,  made  up  of  branched  tubes  of  varying  lengths. 
The  ducts  in  all  are  practically  the  same.  They  are  lobulated  and  supported  by  a  connective- 
tissue  framework.  The  nature  of  the  lumen,  and  the  characters  of  the  epithelium  lining 
the  alveoli,  constitute  the  chief  differences  between  them.  In  the  true  salivary  glands  («)  the 
lumen  of  the  alveoli  is  small,  and  they  are  lined  by  a  single  layer  of  lovv-  columnar  epithelial 
cells,  each  with  a  nucleus  near  its  attached  end.  These  cells  are  markedly  'granular,'  and, 
as  Heidenhain  has  shown,  they  vary  in  appearance  during  rest  and  vi'hen  they  are  secreting. 
In  true  mucous  glands  {b)  there  are  two  kinds  of  cells — one,  mucous  cells,  moderate!}-  tall, 
clear  columnar  epithelial  cells,  each  with  a  small  nucleus  pushed  cjuite  out  to  the  membrana 
propria  of  the  alveolus.  They  are  in  fact  not  unlike  goblet-cells,  and  contain  a  fine  network, 
whose  meshes  are  filled  with  mucigen  (Heidenhain)  ;  this  yields  mucin,  which  stains  deeply 
with  logwood.  Outside  these,  at  various  parts  between  them  and  the  al\-eolar  walls,  are  the 
'  crescents '  (Gianuzzi),  or  parietal  granular  cells.  They  are  small  nucleated  polyhedral  cells. 
In  muco-salivary  glands  (c)  are  found  alveoli,  which  present  the  characters  of  true  salivary 
glands,  and  others  exactly  like  those  of  mucous  glands. 

PREPARATION  ((?). —Place  the  sub-maxillary  glands  of  a  dog  and  rabbit  in  absolute 
alcohol  for  forty-eight  hours.  Make  transverse  sections.  Stain  with  logwood,  and  mount 
one  in  dammar  and  another  in  Farrant's  solution.  This  is  the  best  method  for  a  general 
survey. 

(/;)  Harden  another  gland  in  chromic  acid  and  spirit  mixture  for  a  week,  and  stain  sections 
with  logwood  and  mount  in  Farrant's  solution. 

{c)  Small  pieces,  the  size  of  half  a  pea,  may  be  hardened  in  a  quarter  per  cent,  osmic  acid, 
and  small  pieces  teased  in  glycerine,  or  sections  may  be  made  and  mounted  in  glycerine. 
It  brings  out  the  '  crescents '  very  clearly  in  the  mucous  glands. 

Sub-maxillary  Gland  of  a  Dog.  EXAMINATION  (L). — Observe  the  lobulated  character  of 
the  section  ;  the  fibrous  capsule  sending  septa  into  the  gland,  and  containing  blood-vessels, 
lymphatics,  and  nerves.  Notice  the  alveoli  or  gland-substance  cut  in  every  direction,  and  in 
the  interlobular  septa  here  and  there  a  large  duct — lobar — cut  into.  (H).  Study  a  transverse 
section  of  a  large  lobar  duct.  Notice  the  columnar  epithelium  lining  it,  with  vertical  lines  due 
to  the  longitudinal  arrangement  of  the  fibrils,  and  the  nucleus  placed  about  the  middle  of  the 
cell.  Within  a  lobule  may  be  found  sections  of  an  intra-lobitlar  djict,  which  have  a  small 
lumen  lined  by  columnar  epithelium. 

Study  an  Alveolus. — Observe   the  secretory  epithelium  lining  it.     The  epithelium  may  be 


SALIVARY  GLANDS.  65 

quite  clear,  with  its  small  nucleus  near  the  alveolar  wall.     Study  the  lunate  cells  or  crescents 
of  Gianuzzi.     {Indicate  the  alveoli,  the  cells  lining  tkein,  and  a  lobar  duct  in  PL  XIII.,  Fig.  i.) 

Sub-maxillary  Gland  of  a  Eabbit :  EXAMINATION  (L).  — Observe  the  same  general 
characters. 

(H).  Study  an  Alveolus.— Observe  the  small  lumen  and  the  single  layer  of  low  columnar 
cells  lining  it.     Compare  it  with  the  above  description. 


TONSILS. 

The  tonsils  are  folds  of  mucous  membrane  containing  a  large  number  of  lymph-follicles 
like  those  composing  a  Payer's  patch  (p.  69).  The  mucous  membrane  is  folded  so  as  to 
eave  pits  or  depressions  visible  on  the  surface.  The  epithelium  covering  them  is  stratified, 
and  under  it  is  a  connective-tissue  mucosa,  which  contains  the  closely  aggregated  masses  of 
adenoid  tissue. 

PREPARATION. —  Harden  the  tonsils  for  two  weeks  in  chromic  acid  and  spirit  mixture,  or 
in  Muller's  fluid  for  three  weeks.  Make  transverse  sections,  stain  with  logwood,  and  mount 
in  dammar. 

EXAMINATION  (L). — Note  the  stratified  epithelium,  and  under  it  the  oval  or  rounded 
masses  of  adenoid  tissue,  which  sometimes  penetrate  into  the  epithelium. 
(H).  The  ordinary  structure  of  adenoid  tissue. 


CESOPHAGUS. 

PREPARATION. — Slightly  distend  the  oesophagus  of  a  dog  with  the  chromic  acid  and  spirit 
mixture,  and  keep  it  distended  by  placing  a  ligature  on  its  upper  and  lower  ends.  Suspend  it 
in  a  large  quantity  of  the  same  fluid  until  it  is  '  fixed  ' — i.e.  for  twenty-four  hours — then  cut  it 
into  pieces  one  inch  long  and  continue  the  hardening  for  a  week  or  ten  days.  Make  transverse 
sections  through  the  upper  and  lower  parts  of  the  cesophagus.  Stain  them  with  picrocar- 
mine,  and  mount  in  Farrant's  solution. 

Transverse  section  of  the  (Esophagus.  EXAMINATION  (L). — Observe  the  mucous  coat 
lined  b\'  stratified  epithelium  with  well-marked  mucous  glands  opening  on  its  surface.  The 
mucous  membrane  is  beset  with  small  papillae.  In  the  deeper  part  of  the  mucous  coat  observe 
the  cut  ends  of  the  muscularis  mucosae,  composed  of  non-striped  muscular  tissue  arranged  in 
bundles.  In  the  sub-mucous  coat  observe  the  alveoli  of  the  mucous  glands  and  trace  a  duct 
to  the  surface.  Observe  the  muscular  coat — the  outer  or  longitudinal  layer — cut  transversely, 
and  the  circular  fibi'es  inside  it.  These  contain  much  striped  muscle  in  the  upper  part  of  the 
oesophagus. 

(H).  Study  the  stratified  epithelium  and  the  mucous  glands,  which  are  exactly  like  those 
already  described  in  the  mouth  (p.  61).  A  preparation  mounted  in  dammar  shows  their 
'  mucous  '  characters  admirably. 

Nerves  of  the  (Esophagus. — The  nerve-terminations  are  best  studied  in  the  cesophagus  of  a 
rabbit  by  means  of  the  lemon-juice  and  gold  method.  The  process  is  described  at  p.  71,  in 
connection  with  the  small  intestine. 

K 


66  PRACTICAL   HISTOLOGY. 


STOMACH. 

PREPARATION. — Open  the  stomach  of  a  cat,  dog,  or  rabbit,  and  wash  away  the  layer  of 
mucus  lining  it  with  a  stream  of  salt  solution. 

(fl)  Harden  small  pieces  of  the  pyloric  and  cardiac  ends  and  middle  of  the  stomach  for 
two  weeks  in  the  chromic  acid  and  spirit  fluid  (p.  xxxi).  Complete  the  hardening  in  spirit. 
Make  vertical  sections  of  each  piece.  Stain  sections  with  logwood,  and  mount  them  in 
dammar;  and  others  with  picrocarmine,  and  mount  them  in  Farrant's  solution. 

(/')  After  washing  out  a  stomach  of  any  of  the  above  animals  with  salt  solution,  distend  it 
with  and  place  it  in  absolute  alcohol — or  merely  small  pieces  may  be  hardened  in  absolute 
alcohol.     These  show  gland-structure  well,  and  stain  readily. 

ic)  Very  small  pieces,  not  larger  than  a  pea,  are  to  be  hardened  in  a  quarter  per  cent, 
osmic  acid  for  twenty-four  hours.     Make  sections  and  mount  them  in  Farrant's  solution. 


CARDIAC    END    OF    STOMACH. 

Vertical  Section.  (Logwood  and  dammar.)  EXAMINATION  (L). — Observe  the  mucous 
coat  with  its  gastric  glands  placed  vertically.  Trace  a  gland  and  note  whether  it  branches 
below  or  not.  The  bases  of  the  gland  rest  on  a  small  amount  of  connective  tissue,  which 
sends  up  processes  between  the  glands.  Immediately  outside  this  lies  the  muscularis  mucosae, 
which  consists  of  two  layers — longitudinal  and  circular — and  sends  processes  upwards  be- 
tween the  glands.     {Indicate  this  general  arrangcincni  in  PI.  XIII.,  Fig.  2.) 

The  sub-mucous  coat  composed  of  connective  tissue  containing  blood  and  lymph-vessels, 
fat-cells,  and  nerves.  The  muscular  coat,  consisting  of  two  coats,  with  perhaps  a  third.  The 
appearance  of  these  coats  varies  according  to  the  plane  of  their  section  ;  and  outside  this,  the 
serous  layer,  or  peritoneum.  If  the  stomach  of  the  cat  be  selected,  a  clear  band  of  condensed 
connective  tissue  will  be  observed  lying  immediately  above  the  muscularis  mucosae.  This 
preparation  shows  well  the  passage  of  the  muscularis  mucosa;  fibres  upwards  between  the 
glands. 

To  see  the  finer  details  of  structure  study  the  preparation  mounted  in  Farrant's  solution. 
(L).  Observe  the  same  general  arrangement  as  before.  (H).  Study  z.  peptic  gland.  Note  the 
clear,  tall,  narrow,  columnar  epithelium  lining  the  stomach,  and  continued  some  distance  into 
it.  The  nuclei  of  the  cells  are  placed  near  their  attached  ends.  The  cells  appear  in  some 
cases  to  be  open  at  their  free  ends,  especially  if  the  animal  be  killed  during  digestion,  and 
are,  in  fact,  mucus-secreting  goblet-cells.  Note  that  the  lower  part  of  the  cell  is  always 
granular,  even  though  the  upper  two-thirds  be  quite  transparent.  In  the  middle  and  lower 
part  of  the  gland  note  the  coarsely  granular  nucleated  outer,  ovoid,  or  peptic  cells,  which 
frequently  cause  a  bulging  on  the  side  of  the  gland  ;  and,  internal  to  these,  small  ill-defined 
inner  or  central  cells,  best  seen  at  the  lower  part  of  the  gland.  A  small  narrow  lumen  may  be 
detected  in  the  gland.  The  cells  most  deeply  stained  are  the  ovoid  cells.  Study  the  mus- 
cularis mucosae,  and  note  that  it  consists  of  two — sometimes  three — layers  of  non-striped 
muscle,  and  trace  processes  from  it  upwards  between  the  gland. 

Transverse  sections  of  the  glands  are  to  be  met  with  in  the  same  section.  They  are 
usually  in   groups  of  four  or  five,  and  separated  by  a  little  connective  tissue  and  a  few  non- 


Pla:te  xm     Sub-maxillary  Gland  &,  Stomach 


Minterx--  Bros  litK 


STOMACH.  ■  67 

striped  muscular  fibres.  Observe  the  relation  of  the  outer  and  inner  cells,  and  notice 
the  small  lumen  of  the  gland.  {Indicate  the  appearance  of  transverse  sections  of  the  tubes  in 
PI.  XIII..  Fig.  4.) 

Aniline  Blue.  Stain  a  section  with  a  watery  solution  of  aniline  blue.  Mount  it  in  glycerine 
or  dammar.     The  peptic  cells  are  most  deeply  stained. 

PYLORIC   END    OF    STOMACH. 

EXAMINATION.— Observe  the  mucous  coat,  containing  glands  (mucous)  which  branch 
frequently,  and  do  not  contain  any  large  ovoid  cells.  The  gland-ducts  are  relatively  long, 
and  two  or  three  tubes  open  into  a  short  neck.  The  gland-tube  is  lined  by  the  same  kind  of 
epithelium  throughout.  Notice  transverse  and  oblique  sections  of  the  gland  at  the  lower 
part  of  the  mucous  membrane.  Here  and  there,  near  the  base  of  the  gland,  detached 
patches  of  adenoid  tissue  may  be  seen.     {Indicate  the  general  arrangement  in  PL  XIII.,  Fig.  5.) 

Take  another  section,  stain  it  with  carmine,  and  mount  in  Farrant's  solution.  Notice  the 
same  general  arrangement  as  above.  Observe  that  the  glands  are  lined  throughout  with 
columnar  epithelium.  Study  a  transverse  section,  and  note  the  basement-membrane  of  a 
gland.     Notice  a  few  leucocytes  lying  near  the  bases  of  the  glands. 

Isolation  of  the  Gastric  Glands.  PREPARATION. —  Place  small  pieces  of  the  mucous 
membrane  of  the  stomach  of  a  rabbit  in  five  per  cent,  ammonium  chromate  for  three  days. 
Wash  away  the  colouring  matter  from  a  small  piece  of  the  membrane,  and  place  it  in  picro- 
carmine  solution  for  twenty-four  hours  ;  tease  a  small  piece  in  glycerine.  This  method  shows 
the  shape  and  general  characters  of  the  glands,  and  their  cellular  contents,  the  ovoid  cells 
being  specially  distinct.  The  nucleus  is  stained  of  a  bright  red,  and  a  beautiful  intra-cellular 
plexus  of  fibrils  is  revealed.  Note  also  the  basement-membranes  of  the  gland-tubes.  {Indicate 
the  cells  in  the  gland-tube  in  PI.  XIII.,  Fig.  3.) 

The  osinic  acid  preparations  show  the  peptic  cells  very  well.  They  are  more  deeply 
stained  than  the  other  cells,  and  hence  they  stand  out  more  clearly. 

Blood-vessels  of  the  Stomacli.  PREPARATION. — Inject  the  blood-vessels  of  the  stomach  of  a 
cat  or  dog  through  the  gastro-duodenal  artery  with  a  carmine  gelatine  mass  (p.  li),  and  place 
the  organ  in  alcohol.  Make  vertical  sections  and  stain  them  with  logwood  and  mount  in 
dammar.     If  a  blue  mass  be  used  stain  the  sections  with  carmine. 

EXAMINATION  (L). — Observe  the  vascularity  of  the  mucous  membrane  ;  sections  of  the 
large  vessels  will  be  found  in  the  submucous  coat ;  and  from  these  capillaries  proceed  upwards 
between  the  glands,  and  form  a  capillary  plexus  round  them.  The  veins  rise  near  the  surface, 
and  run  downwards  towards  the  submucous  coat. 


DUODENUM.     SMALL    INTESTINE. 

PREPARATION. — Wash  out  the  contents  of  the  duodenum  of  a  dog  or  cat  with  a  stream 
of  salt  solution,  and  place  pieces  one  inch  long  in  chromic  and  spirit  fluid  for  two  weeks.  If 
the  duodenum  of  a  pig  can  be  obtained,  it  is  to  be  preferred.  Place  similar  pieces  of  the  small 
intestine  some  with,  others  without  a  Peyer's  patch,  in  the  same  hardening  fluid  for  the  same 
time.     Complete  the  hardening  in  alcohol.     Make  transverse  sections.     Stain  a  section  of  the 


68  PRACTICAL   HISTOLOGY. 

duodenum  with  logwood,  and  mount  it  in  dammar.  Stain  sections  of  the  intestine — with  and 
without  a  Peyer's  patch — with  logwood  ;  mount  one  in  dammar,  and  another  in  Farrant's 
solution.  Place  sections  of  the  small  intestine  in  a  quarter  per  cent,  .solution  of  osmic  acid 
for  twenty-four  hours,  and  mount  a  very  thin  section  in  Farrant's  .solution. 

Vertical  Section  of  the  Stomach  and  Duodenum. — Make  a  vertical  section  through  the  pyloric 
valve,  so  as  to  include  a  part  of  the  stomach  and  the  upper  part  of  the  duodenum.  Stain  a 
.section  in  logwood,  and  mount  it  in  dammar. 

EXAMINATION  (L). — Ob.serve  the  gastric  mucous  membrane  with  its  mucous  glands,  and 
trace  a  gradual  transition  from  these  into  the  compound  tubular  and  more  branched  glands 
of  the  duodenum — Brunner's  glands,  which  present  exactly  the  same  structure  as  the  mucous 
glands  of  the  stomach,  with  this  difference,  that  their  duct  is  larger,  and  each  duct  has  a 
greater  number  of  long  branched  tubes  attached  to  it.  The  alveoli  lie  in  the  submucous 
layer  outside  the  muscularis  mucosie,  and  from  these  the  duct  lined  by  columnar  cells  pierces 
the  muscularis  mucosa;,  and  ascends  between  the  crypts  of  Lieberkiihn  to  open  on  the  surface. 
A  few  villi  are  seen  on  the  duodenal  mucous  membrane.  At  the  line  of  junction  of  the 
mucous  membrane  of  the  stomach  and  duodenum  the  muscularis  mucosae  is  absent. 


SMALL   INTESTINE. 

Transverse  Section.  (Logwood  and  dammar.)  EXAMINATION  (L). — Mucous  coat.— 
Observe  the  conical  pointed  villi  ;  .some  of  them  may  be  contracted  and  show  the  epithelial 
covering  thrown  into  folds.  Each  villus  consists  of  a  central  core  covered  with  columnar 
epithelial  cells,  with  their  nuclei  placed  near  the  attached  end  of  each  cell.  Beneath  the  villi 
the  mass  of  the  mucous  membrane  is  made  up  of  Liebcrkuhns  follicles,  consisting  of  simple 
tubular  glands  lined  by  epithelium.  Note  the  adenoid  tissue  between  the  bases  of  the 
glands,  and  outside  this  the  muscularis  viucosce.  The  subunicous  coat  consists  of  connective 
ti.ssue  containing  blood-vessels  and  nerves.  Muscular  coat. — Note  the  thick  inner  circular 
layer,  which  will  appear  different  according  as  the  section  has  been  made  across  or  parallel  to 
the  long  axis  of  the  gut ;  if  the  latter,  note  that  the  muscular  fibres  are  arranged  in  blocks 
separated  and  surrounded  by  connective  tissue  ;  outside  this  the  narrower  longitudinal  layer, 
and  outside  all  the  serous  coat.  This  specimen  does  best  for  a  general  study  of  the  relation  of 
parts.  Select  a  villus  and  examine  it  with  (H).  Notice  the  long  fusiform  nuclei  in  that  part 
of  the  core  of  the  villus,  next  the  central  lacteal  vessel.  They  are  the  nuclei  of  non-striped 
muscular  fibres,  which  pass  from  the  muscularis  mucosae  into  the  villi.  (^Indicate  the  general 
arrangeincHt  in  one-half  of  Y\.  XIV.,  Fig.  i.) 

Examine  now  a  similar  preparation  stained  with  logwood,  but  mounted  in  Farrant's  solu- 
tion. Observe  the  same  relation  of  parts  as  described  above.  Fix  a  villus  in  the  field,  and 
observe  (H)  the  epithelial  investment,  consisting  of  columnar  nucleated  cells  with  a  clear  hem 
containing  vertical  striae  (compare  p.  lO,  on  epithelium).  Observe  goblet-cells  scattered  between 
the  epithelial  cells  and  study  them.  [Indicate  the  covering  and  structure  of  a  villus  in  PI. 
XIV.,  Fig.  2).  Search  for  these  cells  seen  en  face.  In  the  substance  of  the  villus  observe  the 
adenoid  tissue  of  which  it  consists,  with  nuclei  in  its  meshes,  and  perhaps  a  lacteal  vessel  in 
its  centre. 

Lieberkiiiin's  Glands.  Study  one.  Observe  its  test-tube  shape,  and  the  low  nucleated 
columnar  epithelium  lining  it.  At  their  bases  and  outside  them  observe  adenoid  tissue. 
Study  the  two  layers  of  the  muscularis  mucosa;. 


SMALL   INTESTINE.  bg 

Submucous  coat. — Observe  its  blood-vessels,  and  perhaps  nerve-ganglia  {Meissners  plexus), 
and  perhaps  a  solitary  gland. 

Muscular  Coat. — Observe  the  characters  of  non-striped  muscle  cut  transversely  and  longi- 
tudinally. 

PEYER'S    PATCH. 

Transverse  Section.  (Logwood  and  dammar.)  EXAMINATION  (L). — Observe  the  masses 
of  adenoid  tissue  in  the  submucous  coat.  Their  conical  points  project  upwards  into  the  gut, 
and  are  covered  by  epithelium,  but  no  villi  e.\;ist  over  them. 

Treble-staining  of  the  Small  Intestine.  PREPARATION. — Take  one  of  the  transverse  sections 
of  the  small  intestine,  containing  a  Peyer's  patch.  Stain  it  with  picrocarmine,  wash  it  lightly 
in  water  slightly  acidulated  with  acetic  acid.  Stain  it  now  with  solution  of  iodine  green 
(p.  xlv)  until  it  becomes  slightly  green,  wash  it  quickly  in  water,  and  mount  it  in  dammar, 
taking  care  that  it  does  not  remain  too  long  in  the  alcohol. 

EXAMINATION  (L). — This  is  an  exquisitely  beautiful  preparation.  All  the  glands,  and 
especially  Peyer's  patches,  are  green,  all  connective  tissue  is  red,  and  so  the  submucous  layer 
stands  out  bright  red.  The  muscularis  mucosa;  and  muscular  coats  are  yellow.  Observe 
the  interruption  of  the  muscularis  mucosa,  where  the  adenoid  follicles  reach  the  surface.  A 
thin  layer  of  connective  tissue  lies  superficial  to  the  muscularis  mucosae.  Such  preparations 
keep  a  long  time. 

In  the  preparations  steeped  in  osmic  acid,  after  hardening  with  the  chromic  acid  mixture, 
the  various  details  are  seen  with  exquisite  clearness. 

Blood-vessels  of  the  Small  Intestine. — Make  transverse  sections  of  the  small  intestine  of  a 
cat  or  dog  whose  blood-vessels  have  been  filled  with  a  carmine  and  gelatine  mass.  Mount  in 
dammar.     It  is  easy  to  inject  the  intestine  from  the  superior  mesenteric  artery. 

EXAMINATION  (L). — Observe  the  villi,  and  note  their  vascularity.  Usually  one  artery 
ascends  on  one  side,  and  splits  into  capillaries,  which  are  distributed  closely  under  the  epithe- 
lium. The  vein  descends  on  the  opposite  side  of  the  villus.  Note  the  rich  plexus  of  capillaries 
surrounding  Lieberkiihn's  glands,  and  the  large  arterial  trunks  in  the  submucous  coat.  Study 
the  distribution  of  the  blood-vessels  in  the  muscular  coat,  and  observe  that  it  is  not  so  vascular 
as  the  mucous  coat.  {Indicate  the  arrangement  of  the  blood-vessels  in  one  half  of  Vl.  'K.YV ., 
Fig.  I.) 

Injected  Villi  seen  from  above. — Mount  in  dammar  a  small  piece  of  the  small  intestine  of  a 
rabbit  whose  blood-vessels  have  been  injected  with  a  carmine  and  gelatine  mass.  Place  the 
section  in  the  slide  so  that  the  villous  surface  is  uppermost. 

EXAMINATION  (L).— Observe  the  leaflet-like  injected  villi  directed  towards  the  observer, 
and  by  focussing  through  the  thickness  of  the  preparation  the  course  and  distribution  of  the 
larger  arterial  trunks  in  the  submucous  coat  can  be  clearly  made  out. 

Lymphatics  or  Lacteals  of  the  Small  Intestine. — The  origin  of  the  lacteal  within  the  central 
part  of  a  villus  as  a  blind  tube  has  already  been  noted  (p.  6'&),  but  the  branches  of  the 
lymphatics  in  the  walls  of  the  gut  can  easily  be  studied,  after  injection  with  Berlin-blue,  by  the 
puncture  method  ;  an  excellent  view  of  their  arrangement  will  be  obtained  in  the  prepara- 
tion of  the  intestine  treated  with  silver  nitrate  (p.  37),  to  show  the  cement-substance  of 
muscle.     This  will  be  referred  to  again  under  lymphatics  (p.  76). 


70  PRACTICAL   HISTOLOGY. 


VERMIFORM    APPENDIX. 

PREPARATION.—  Inject  the  walls  of  the  vermiform  appendix  of  a  rabbit  with  a  two 
per  cent,  watery  solution  of  Berlin-blue,  which  readily  passes  into  the  lymphatics.  Harden  in 
alcohol,  and  make  transverse  sections.  Stain  them  with  picrocarmine,  and  mount  in  Farrant's 
solution. 

EXAMINATION  (L). — Observe  the  masses  of  adenoid  tissue,  stained  red,  divided  into  an 
inner  and  an  outer  set,  and  round  each  the  lymph-paths  filled  with  Berlin-blue.  The  masses 
of  adenoid  tissue  are  exactly  like  Payer's  patches,  and  each  arc  is  partially  surrounded  by  a 
lymph-stream,  as  indicated  by  the  blue  injection,  none  of  which  passes  into  the  interior.  This 
space  represents  a  lympli-simis,  so  that  the  masses  of  adenoid  tissue  are  partially  suspended 
in  a  lymph-stream.  Exactly  the  same  relation  obtains  in  a  Payer's  patch.  {^Indicate  these 
appearances  in  PI.  XIV.,  Fig.  5.) 


LARGE    INTESTINE. 

PREPARATION. — After  washing  out  the  large  intestine  of  a  dog  or  cat,  cut  it  into  pieces 
one  inch  square  and  harden  them  in  chromic  acid  and  spirit  fluid  for  two  weeks  and  complete 
the  hardening  in  spirit.  Make  transverse  sections.  Stain  with  logwood,  and  mount  one  in 
dammar,  and  another  in  Farrant's  solution. 

Vertical  Section  of  the  Large  Intestine.  (Logwood  and  dammar.)  EXAMINATION  (L). — 
Observe  the  mucous  coat  thrown  into  folds,  owing  to  the  contraction  of  the  muscular  coat.  It 
is  devoid  of  villi.  Observe  the  vertically  set  Lieberkiihn's  glands,  exactly  like  those  in  the 
small  intestine.  {Indicate  one  in  PI.  XIV.,  Fig.  3.)  Some  of  them  are  sure  to  be  cut  trans- 
versely, when  they  present  a  honeycomb-like  appearance.  (Indicate  this  in  PI.  XIV.,  Fig.  4.) 
Observe  the  sub-mucous  and  muscular  coats,  very  like  those  of  the  small  intestine.  Per- 
haps a  solitary  gland  may  be  found  in  the  sub-mucous  coat  with  its  apex  projecting  free  into 
the  mucous  coat.  There  is  a  well-marked  muscularis  mucosae  in  the  mucous  coat,  which 
is  pierced  by  the  solitary  glands. 

In  the  preparation  mounted  in  Farrant's  solution,  examine  (H)  the  glands,  both  when 
divided  vertically  and  transversely,  and  observe  the  adenoid  tissue  laden  with  lymph-corpuscles, 
between  and  supporting  them. 

Solitary  glands  will  be  found  in  sections  of  the  large  intestine.  They  are  simply  masses 
of  adenoid  tissue  placed  in  the  sub-mucous  coat.  They  are  easily  stained  by  the  same 
methods  as  directed  for  Payer's  patches  (p.  69). 

Blood-vessels  of  the  Large  Intestine  are  prepared  exactly  in  the  same  way  as  those  of  the 
small  intestine.  Several  years  ago  it  was  customary  to  use  an  opaque  injection,  which 
gives  a  beautiful  effect  when  the  piece  of  intestine  is  mounted  upon  a  dark  ground  with  the 
mucous  surface  uppermost,  and  illuminated  by  means  of  a  condenser.  The  mouth  of  each  gland 
is  surrounded  by  a  plexus  of  capillaries,  so  that  the  injection  presents  a  regular  honeycomb- 
like appearance. 

Similar  preparations  made  of  the  gastric  mucous  membrane  at  first  sight  resemble  that  of 
the  large  intestine.  The  gastric  mucosa  is  recognised  by  the  existence  of  small  openings 
at  the  bottom  of  the  depressions,  and  the  latter  are  not  so  regular  as  those  in  the  large 
intestine. 


^=^LATE3av:  Small  &  Lj\rge  iNTEST^ffi 


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1 

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"MitM.ei-n.  Bros  1il1"i 


LARGE  INTESTINE.  71 


NERVE-PLEXUSES    OF    THE    INTESTINE. 

There  are  two  plexuses  of  nerves,  consisting  chiefly  of  non-medullated  nerve-fibres  and 
ganglionic  cells.  In  the  intestinal  walls,  one — plexiis  myentericus  of  Meissner — lies  in  the 
submucous  coat;  the  other — AuerbaMs plexus — lies  between  the  muscular  coats. 

PREPARATION. — Auerbach's  plexus  is  best  prepared  in  the  following  way.  ia)  Wash  out  a 
loop  of  perfectly  fresh  small  intestine  of  a  rabbit,  and  distend  it  with  the  juice  of  a  fresh  lemon  ; 
ligature  both  ends  of  the  gut  and  place  it,  still  distended,  in  lemon-juice  for  from  five  to  seven 
minutes.  Open  the  ligatures  and  wash  it  thoroughly  in  water,  and  fill  it  with  a  two  per  cent,  gold 
chloride  solution  ;  ligature  the  gut  as  before,  and  suspend  it  for  half  an  hour  in  a  one  percent. 
gold  solution.  Wash  it  thoroughly,  and  transfer  it  to  a  twenty-four  per  cent,  dilution  of  formic 
acid,  to  reduce  the  gold.  The  preparation  must  be  kept  in  the  dark.  After  reduction,  it  has 
a  beautiful,  rich,  reddish-brown  colour.  Wash  it  thoroughly  and  keep  it  in  a  preservative 
fluid  (p.  xl).  With  forceps  it  is  easy  to  peel  off  strips  of  the  outer  muscular  layer,  to  which 
Auerbach's  ple.xus  adheres.  Mount  in  glycerine.  The  separation  takes  place  more  easily 
after  maceration  for  a  week  in  the  preservative  fluid.     This  is  the  best  method. 

(3)  Another  method  is  to  use  dilute  alcohol,  as  the  distending  and  softening  medium,  for 
forty-eight  hours  ;  then  to  peel  off  the  outer  muscular  coat  as  directed  above.  Stain  it  with 
logwood,  and  mount  in  glycerine.     This  gives  fairly  good  results. 

[c]  One-twentieth  per  cent,  acetic  acid  may  be  used  in  the  same  way  to  fill  the  gut  and  to 
macerate  it,  for  thirty  hours.  After  maceration  and  washing  it  is  well  to  steep  the  gut  for  a 
short  time  in  five  per  cent,  solution  of  sodic  bicarbonate,  to  get  rid  of  the  acid.  Peel  off  the 
muscular  coat  as  before,  stain  it  v\'ith  logwood  and  mount  it  in  glycerine. 

EXAMINATION  (L). — Observe  the  v/ide-meshed  plexus  of  fairly  regular  form.  At  the 
nodes,  groups  of  ganglionic  cells  are  to  be  detected.  Ganglionic  cells  are  either  in  groups,  or 
sometimes  disposed  lengthwise  in  the  flat,  band-like  nerve-branches.  Each  band  has  an 
endothelial  sheath.  (H).  The  cells  are  very  variable  in  size,  and  some  of  them  are  multipolar. 
Nerve-branches  are  given  off  from  this  plexus  to  supply  the  longitudinal  and  circular  muscular 
coats,  but  their  mode  of  termination  is  unknown. 

The  Plexus  of  Meissner  is  prepared  in  a  similar  way,  and  it  supplies  the  muscularis  mucosae, 
the  blood-vessels  and  glands  of  the  mucous  coat.  Its  meshes  are  much  wider  and  less 
regular  than  Auerbach's.     The  nerve-cells  are  easily  seen  in  it. 

In  vertical  sections  of  a  cat's  small  intestine,  hardened  in  chromic  acid  and  spirit,  groups 
of  ganglionic  cells  may  easily  be  detected  in  the  sub-mucous  coat.  These  are  the  ganglia  of 
Meissner's  plexus. 


THE    LIVER. 

It  is  well  to  examine  the  liver  of  several  animals  as  they  differ  in  the  complexity  of 
structure.  It  is  best  to  begin  with  a  pig's  liver,  and  to  pass  to  that  of  a  rabbit,  a  dog  or  cat, 
and  then  to  the  human  organ.     The  liver  may  be  hardened  in  several  ways. 

PREPARATION.  A.  Chromic  Acid  and  Spirit  Mixture. — Place  pieces  of  a  perfectly  fresh 
liver  (pig,  rabbit,  cat  or  dog,  and  man),  about  half  an  inch  square,  in  a  large  quantity  of  the 
above  fluid.  Do  not  wash  them.  As  the  liver  is  very  vascular,  much  blood  exudes  and 
causes  a  precipitate  in  the  fluid,  which  must  be  changed  at  the  end  of  twelve  hours.     As  soon 


72  PRACTICAL    HISTOLOGY. 

as  the  fluid  appears  turbid  it  ought  to  be  changed.     The  hardening  will  be  completed  in  two 
to  three  weeks.     Wash  the  tissues  well,  and  transfer  them  to  spirit  until  they  are  required. 

B.  Miiller's  Fluid.—  Larger  pieces  of  the  liver  may  be  hardened  in  this  way,  as  this  fluid 
penetrates  organs  more  readily  than  A.  It  takes,  however,  longer  to  harden  (four  to  six 
weeks). 

C.  Two  per  cent.  Potassic  Bichromate,  which  acts  in  the  same  way,  and  in  the  same  time,  as 
Miiller's  fluid. 

Make  sections  of  the  liver  of  pig,  rabbit,  cat  or  dog,  and  man,  by  means  of  a  freezing 
microtome,  and  see  that  the  sections  are  thin  and  made  through  the  fibrous  capsule.  Some  of 
them  are  to  be  stained,  and,  for  non-stained  preparations,  place  the  sections  in  one  per  cent, 
osmic  acid  for  four  or  five  hours.  This  is  an  excellent  method,  and  sharpens  the  outlines  of 
all  the  tissues. 

I.     LIVER    OF   A    PIG. 

EXAMINATION  (L). — Stain  a  section  with  logwood,  and  mount  it  in  dammar.  Observe 
the  fibrous  capsule,  and  note  the  processes  it  sends  into  the  organ,  where  they  become 
continuous  with  the  fibrous  sheath  which  surrounds  each  lobule.  Note  that  the  section  is 
mapped  out  into  a  number  of  distinct,  more  or  less  polygonal  areas  of  nearly  equal  size 
by  bands  of  connective  tissue  stained  blue  ;  these  areas  are  the  lobules,  and  each  one  is  com- 
pletely invested  by  a  fibrous  sheath — a  continuation  of  Glisson's  capsule.  Within  each  of  these 
areas  is  the  true  gland-substance,  which  will  be  considered  in  other  preparations.  This  pre- 
paration gives  the  student  an  excellent  idea  of  the  lobular  character  of  the  liver.  {Indicate  the 
lobules  ill  PI.  XV.,  Fig.  i.) 

2.     LIVER  OF   A    RABBIT. 

Logwood  and  dammar.  Note  that  the  liver-lobules  are  not  nearly  so  accurately  mapped 
off  from  each  other,  but  still  polygonal  areas— sections  of  the  lobules — can  be  seen  mapped 
off  here  and  there  from  each  other  by  a  very  small  quantity  of  connective  tissue — interlobular 
comiective  tissue,  or  Glisson's  capsulc^s\.d.med  blue.  {Indicate  these  appearances  in  PI.  XV., 
Fig.  2.) 

3.    THE    LIVER    OF   A    DOG   AND    OF   MAN. 

These  ought  to  be  prepared  and  studied  in  the  same  way.  In  them  the  outlines  of  the 
lobules  are  even  less  distinct,  owing  to  the  lobules  being  more  or  less  confluent,  due  to  the  small 
amount  of  interlobular  connective  tissue.  It  is,  therefore,  the  amount  of  interlobular  con- 
nective tissue  which  determines  the  mapping  out  of  one  lobule  from  another.  Note 
particularly  the  thickness  of  the  fibrous  capsule,  and  the  small  amount  of  interlobular 
connective  tissue  in  a  normal  human  liver.  This  is  essential,  as  in  some  diseases — e.g.  cirrhosis 
— it  is  increased  in  amount,  and  it  is  necessary  to  accustom  the  eye  to  the  normal  amount. 
Continue  the  examination  of  the  human  liver  with  (L).  Observe  the  capsule,  and  note  its 
thickness,  and  that  it  consists  of  two  layers  ;  the  lower  one  sends  very  fine  processes  into 
the  organ  between  the  lobules.  These  processes  are  better  seen  in  a  section  where  the 
liver-cells  have  been  pencilled  or  shaken  out,  as  directed  for  adenoid  tissue  (p.  29).  Study  a 
lobule.  In  its  centre,  if  it  be  cut  transversely,  observe  the  hepatic  or  intra-lobularvein,  and  note 
its  size  ;  for  it  is  sometimes  dilated  in  diseased  conditions,  especially  where  there  is  congestion 
of  the  lungs  and  right  side  of  the  heart.  Radiating  outwards  from  this,  observe  the  columns 
of  nucleated  liver-cells,  united  at  the  outer  part  of  the  lobule  by  transverse  branches  to  form  a 


Plate  .Cv'  li'/er 


Muite;  n  oros  lith 


THE   LIVER.  73 

network,  [Indicate  the  columns  of  liver-cells  in  PL  XV.,  Fig.  3.)  The  radiate  arrangement  is 
best  seen  in  the  columns  of  cells  nearest  the  hepatic  vein.  Between  these  cell-columns  narrow 
spaces  exist,  which  indicate  the  position  of  the  intra-lobular  blood-capillaries.  In  an  unin- 
jected  liver  the  capillaries  appear  relatively  small.  Search  between  lobules  for  a  transverse  section 
of  a  branch  of  the  portal  or  interlobular  vein,  hepatic  artery,  and  bile-duct,  all  of  which  run  to- 
gether and  are  surrounded  by  connective  tissue,  which  constitutes  part  of  Glisson's  capsule. 
[Indicate  these  structures  in  PI.  XVI.,  Fig.  3.)  The  channels  in  which  they  lie  are  known  as 
portal  canals.  Fix  the  bile-duct  in  a  field  and  examine  with  (H).  The  bile-duct  may  be  easily 
distinguished  on  account  of  its  being  lined  by  low,  columnar,  almost  cubical,  epithelium.  In  its 
wall  may  be  found  circularly  disposed  non-striped  muscular  fibres,  and  if  a  particularly  large 
bile-duct  be  found,  the  walls  may  be  seen  to  contain  sections  of  mucous  glands,  which  secrete 
mucin  and  add  it  to  the  bile  as  it  passes  along  the  bile-ducts.  The  portal  vein  is  large,  and 
neither  it  nor  the  hepatic  artery  present  anything  peculiar  in  their  characters.  Surrounding 
these  is  the  connective  tissue  of  the  capsule  of  Glisson.  The  tissue  is  more  or  less  lamellar  ; 
the  bundles  run  in  various  directions,  and  between  them  are  flattened  connective-tissue  cor- 
puscles stained  blue.  Note  its  amount.  A  very  small  amount  of  intra-lobular  connective  tissue 
is  to  be  found  ;  it  is  best  seen  around  the  hepatic  veinlet. 

Study  the  liver-cells  (H). — They  are  polygonal  or  cubical  cells  of  nearly  the  same  size, 
uniformly  granular  in  appearance,  are  devoid  of  an  envelope,  and  contain  a  spherical  and 
deeply-stained  nucleus,  though  some  may  contain  two  nuclei,  and  rare  examples  are  met  with 
without  a  nucleus.  The  cells  at  the  outer  part  are  usually  slightly  smaller  than  the  others. 
Note  particularly  the  network  formed  by  these  cells,  and  how  they  intertwine  with  the  blood- 
capillaries  between  them.  Each  cell  contains  a  dense  intra-cellular  and  intra-niiclear  plexus 
of  fibrils,  which  give  it  a  granular  appearance  (PI.  XV.  Fig.  4).  See  p.  13,  where  these  points 
are  alluded  to. 

Stain  similar  sections  with  logwood  and  picrocarmine,  and  mount  them  in  Farrant's  solu- 
tion.    In  this  medium  the  finer  details  can  be  more  carefully  studied. 

Mount  an  unstained  section,  which  has  been  steeped  in  osmic  acid,  in  Farrant's  solution. 
This  is  an  excellent  method  for  sharpening  the  outline  of  the  various  parts.  Any  oil-globules 
present  within  the  liver-cells  are  blackened,  osmic  acid  being  an  excellent  test  for  the  presence 
of  fatty  infiltration  or  degeneration. 

BLOOD-VESSELS    OF   THE    LIVER. 

PREPARATION. — The  blood-vessels  of  the  liver  of  a  rabbit  or  other  animal  may  be  in- 
jected either  from  the  portal  or  hepatic  vein,  or  both.  It  is  well  to  examine  a  liver  whose 
vessels  are  injected  either  with  a  red  or  a  blue  carmine  mass,  or  if  desired  the  portal  vein 
may  be  injected  with  a  blue  mass  and  the  hepatic  vein  with  a  red,  ox  vice  versd.  In  injecting 
from  the  portal  vein  the  mass  should  flow  freely  from  the  hepatic  vein,  which  should  then 
be  ligatured,  and  then  the  blood-vessels  will  fill  properly.  One  must  judge  of  the  amount 
to  be  pressed  into  the  vessels  by  the  appearance  and  colour  of  the  liver-lobules.  Compare 
p.  Hi  for  general  directions.  Harden  the  organ  in  two  per  cent,  potassic  bichromate  and  then 
in  alcohol,  or  place  the  liver  at  once  in  alcohol.  Make  sections,  and  mount  them,  stained  or 
unstained,  in  dammar. 

EXAMINATION  (L). — Select  a  lobule  which  has  been  cut  transversely,  and  observe  the 
origin  of  the  hepatic  vein  in  its  centre,  and  at  the  periphery  of  the  lobule  sections  of  the  portal 
vein.      Within  the  lobule,   and  connecting  these   two  vessels,   observe  the   blood-capillaries 

L 


74  PRACTICAL   HISTOLOGY. 

with  a  distinct  radial  arrangement  near  the  hepatic  vein,  thou^li  the  ineshvvork  becomes 
more  polygonal  at  its  outer  part.  Relatively  the  hepatic  cells  appear  smaller  than  in  the  un- 
injected  specimen.  Sections  of  the  hepatic  vein  issuing  from  within  a  lobule  ought  to  be 
looked  for.  A  double  injection  is,  of  course,  very  instructive,  though  the  areas  of  the  respec- 
tive veins  may  not  be  accurately  mapped  out.  (^Indicate  the  general  arrangcvient  of  the  blood- 
vessels in  PI.  XVI.,  Fig.  i.) 

Opaque  Injections. — It  is  well  to  examine  by  reflected  light  a  section  of  a  liver  whose 
blood-vessels  have  been  injected  by  an  opaque  mass — say  red  in  the  hepatic  vein,  and  j-ellow 
in  the  portal. 

BILE-DUCTS. 

PREPARATION. — Inject  the  fresh  and  still  warm  liver  of  a  rabbit  or  guinea-pig  (killed 
by  bleeding)  from  the  bile-duct  with  a  cold,  watery,  fresh-filtered  solution  of  Prussian  blue 
(p.  li).  This  is'  by  no  means  an  easy  task.  The  great  thing  is  to  use  a  constant  and  not 
too  great  pressure  (p.  liii).  Whenever  the  outer  part  of  the  lobules  begins  to  get  blue,  stop. 
It  is  advantageous  to  inject  a  red  gelatine  mass  into  the  portal  vein  after  the  injection  of 
the  bile-ducts  has  been  completed.  Place  the  liver  in  alcohol  for  a  few  hours,  then  cut  it  into 
pieces,  and  put  some  in  alcohol  and  others  in  Miiller's  fluid  for  two  weeks.  Make  sections  in 
the  usual  way  by  freezing. 

EXAMINATION  (L). — In  a  section  of  a  piece  hardened  in  alcohol  and  mounted  in  dammar 
observe  the  larger  interlobular  bile-ducts,  filled  with  a  blue  mass  between  the  lobules  ;  and 
from  these  fine  branches  can  be  seen  to  pass  into  the  lobules,  within  which  they  form  a 
polygonal  meshwork  over  and  between  the  hepatic  cells.  {Indicate  the  hilc-ducts  in  PI.  XVI., 
Fig.  30 

Stain  a  section  hardened  in  Miiller's  fluid  with  picrocarniine,  and  mount  it  in  Farrant's 
solution. 

EXAMINATION  (L). — Note  the  same  arrangement  as  above,  only  the  hepatic  cells  are 
better  preserved,  and  their  nuclei  stained. 

(H).  Select  a  large  interlobular  bile-duct  and  observe  its  lining  of  low  columnar  epithelium, 
and,  it  may  be,  sections  of  a  mucous  gland  in  its  walls.  The  fusiform  nuclei  of  the  non-striped 
muscle  in  the  wall  of  the  bile-duct  are  stained  red.  Trace  now  the  inti-a-lobular  bile-capillm-ies 
or  channels,  and  observe  their  polygonal  shape,  i.e.,  exactly  the  shape  of  the  liver-cells,  the 
plexus  they  form  over  and  between  the  hepatic  cells,  whose  nuclei  are  stained  red.  The  bile- 
capillaries  are  much  smaller  than  the  uninjected  spaces  of  the  blood-capillaries. 

In  section  the  blue  point  indicating  a  section  of  a  bile-capillar)-  is  found  in  the  angle 
where  three  or  more  cells  meet.  They  are  never  found  between  the  liver-cells  and  blood- 
capillaries,  and  are  always  separated  from  the  capillary  blood-stream  by  a  part  of  one  or 
more  liver-cells. 

Isolated  Liver-cells. — Scrape  the  surface  of  a  section  of  a  liver,  and  diffuse  the  scraping  in 
salt  solution  on  a  slide.  Compare  p.  1 3,  where  these  cells  are  described.  Re-examine  the 
osmic  acid  and  picrocarmine  preparations  of  the  liver  of  the  rabbit  and  newt,  and  in  the  latter 
study  the  intra-cellular  plexus. 

Oil-particles  within  Liver-cells. — These  are  very  common  in  stall-fed  animals — e.g.  ox — and 
are  readily  recognised  by  their  highly  refractive  appearance,  and  by  the  action  of  osmic  acid, 
which  blackens  them  in  a  very  short  time. 


Plate  XvT  .  Liver  &  Pancreas 


^^'Iir\tert\  Brosiitk 


/:> 


PANCREAS. 

The  pancreas  in  many  respects  resembles  the  salivary  glands,  though  there  are  important 
differences.  It  is  lobulated,  and  has  a  connective-tissue  stroma,  blood  and  lymphatic 
arrangements  like  the  salivary  glands.  It  is  a  compound  tubular  gland,  and  the  gland-tissue 
has  lobar  and  inter-lobular  ducts,  which  terminate  by  means  of  an  intermediate  piece  in  the 
alveoli,  which  consists  of  wavy,  branched,  and  convoluted  tubes,  each  having  a  basement-mem- 
brane lined  by  a  single  layer  of  columnar  or  cylindrical  cells,  whose  free  ends  are  sometimes 
conical.  The  substance  of  each  cell  shows  a  marked  division  into  two  zones — the  outer 
(next  the  membrana  propria)  is  homogeneous,  and  stains  easily  and  deeply  with  carmine  and 
logwood- — the  inner  is  coarsely  granular,  and  does  not  stain  readily  with  dyes,  and  in  it  lies 
the  spherical  nucleus.  The  lumen  of  the  alveolus  is  small,  and  so  are  the  alveoli  themselves. 
Heidenhain  has  shown  that  the  appearances  and  relative  sizes  of  these  two  zones  alter  during 
digestion — i.e.  during  physiological  activity,  just  like  the  salivary  glands. 

PREPARATION  {a). — Place  small  pieces  of  the  freshly  excised  pancreas  of  a  dog  in 
absolute  alcohol  for  forty-eight  hours,  and  then  make  sections.  Stain  them  with  carmine,  and 
mount  in  Farrant's  solution,  or,  better  still,  with  logwood,  and  mount  in  dammar. 

{U)   Osmic  acid  preparations  are  also  valuable. 

EXAMINATION  (Lj.— Observe  the  lobules,  the  connective  sheath  and  its  septa,  perhaps 
containing  the  section  of  a  lobar  duct.  Study  a  lobule,  and  note  an  inter-lobular  duct  ;  and 
the  alveoli — relatively  small — cut  in  every  direction.  (H).  Study  an  alveolus :  note  the 
secretory  epithelium,  with  its  outer  zone  stained  of  a  logwood  tint,  and  its  inner  half  granular 
and  unstained.  This  appearance  is  quite  characteristic.  Scarcely  any  lumen  is  observable, 
and  ver)-  little  interstitial  matter  separates  one  secretory  cell  from  another.  {Indicate  the 
appearance  of  the  alveoli,  ami  the  epithelium  lining  them,  in  PI.  XVI.,  Fig.  4.) 


76  PRACTICAL   HISTOLOGY. 


THE  LYMPHATIC  SYSTEM. 

The  lymphatic  vessels  consist  of  capillaries  and  trunks  of  variable  size.  The  capillaries  con- 
sist of  single  layers  of  squames  with  characteristic  sinuous  edges,  and  the  larger  vessels  have 
a  structure  resembling  veins  with  three  coats,  but  the  walls  are  very  much  thinner.  They 
have  valves  in  their  course. 

LYMPHATICS    OF   THE    DIAPHRAGM. 

PREPARATION  (L). — Kill  a  rabbit  or  guinea-pig  by  bleeding.  Tie  a  ligature  round  the 
inferior  vena  cava  and  the  gullet,  and  remove  the  abdominal  viscera,  so  as  to  expose  the  under 
surface  of  the  central  tendon  of  the  diaphragm.  With  a  fine  camel-hair  pencil,  brush  away 
the  epithelium  covering  it,  and  then  silver  it  in  the  usual  way  (p.  xlv).  After  exposure  to 
light  mount  a  small  piece  in  dammar. 

EXAMINATION  (L). — Observe  the  network  of  lymphatic  vessels,  some  with  a  small, 
others  with  a  large  lumen.  Trace  the  course  of  a  large  vessel,  and  observe  the  bulgings  in  its 
course  with  narrower  parts  between  (PI.  XVII.,  Fig.  i).  Especially  at  the  division  of  a  vessel 
a  valve  may  be  seen.  (H).  Study  the  character  of  the  epithelium  lining  the  vessels,  and  notice  its 
sinuous  outline. 

Lymphatics  of  the  Intestine. — For  a  large  class,  silvered  lacteals  are  readily  seen  in  the 
preparation  of  silvered  muscular  fibre  obtained  from  the  small  intestine  of  a  rabbit  (p.  37). 
Refer  to  that  preparation,  and  observe  details  in  the  structure  of  these  vessels  similar  to  those 
described  in  the  lymphatics  of  the  diaphragm. 

The  lymphatics  communicate  with  tlu  semis  cavities  by  means  of  small  apertures — stomata — 
guarded  by  small  granular  cells.  The  stomata  are  most  easily  obtained  from  the  septum  of 
the  great  lymph-sacs  of  the  frog,  though  they  may  also  be  obtained  by  silvering  the  under 
surface  of  the  diaphragm  of  a  rabbit.  Behind  the  stomach  and  on  each  side  of  the  vertebral 
column  of  the  frog  there  lies  a  large  lymph-sac,  separated  only  from  the  peritoneal  cavity  by  a 
very  delicate  membrane  or  septum,  which  is  perforated  by  small  apertures — stomata — which 
bring  the  two  cavities  into  direct  communication. 

SEPTUM    CYSTERN^    LYMPHATICS   MAGNS. 

PREPARATION. — Kill  a  frog  and  carefully  remove  the  abdominal  viscera,  taking  care  not 
to  injure  the  fine  septum  above  described.  Pour  distilled  water  over  the  posterior  ab- 
dominal wall  immediately  behind  the  stomach,  when  a  delicate  membrane  will  be  floated 
up  and   brought  into  view.     Pour  over  it  a  half  per  cent,  solution  of  nitrate  of  silver  till  it 


FtAiEXVII-  Lymphatics 


■^finterm.  Bros  litk 


LYMPHATIC  GLANDS.  y-j 

becomes  slightly  white  ;  snip  out  the  membrane,  and  after  washing  expose  it  to  light  in  the 
usual  way.     Mount  a  small  piece  in  glycerine. 

EXAMINATION  (L  and  H). — Observe  the  membrane  perforated  here  and  there  by  small 
apertures — stoinata — easily  recognised  by  their  having  a  brownish  margin.  Their  margins  are 
bounded  by  small,  plastic,  granular  cells,  stained  brownish  with  the  silver  (PI.  XVII.,  Fig.  2). 
The  stomata  may  either  be  open  or  closed.  In  addition,  silver  outlines  are  seen,  and  by  focussing 
through  the  thickness  of  the  membrane  it  is  seen  that  epithelium  exists  on  both  surfaces,  and 
that  the  size  and  shape  of  the  cells  differ  on  the  two  sides.  On  the  peritoneal  surface  the  cells 
are  elongated,  while  on  the  side  directed  towards  the  lymph-sac  they  are  more  or  less  polygonal 
and  much  broader.  Similar  stomata  exist  on  the  mesogastrium  and  mesentery  of  the  frog, 
and  in  the  intercostal  pleura,  &c.,  of  mammals.     They  are  prepared  in  the  same  way. 


LYMPHATIC   GLANDS. 

(A.)     COMPOUND    LYMPHATIC    GLANDS. 

Lymphatic  glands  are  accumulations  of  adenoid  tissue  (p.  29)  in  the  course  of  the  lym- 
phatic vessels,  and  so  they  have  afferent  and  efferent  vessels.  Each  gland  is  surrounded 
by  a  connective-tissue  capsule,  which  not  unfrequently  (ox)  contains  non-striped  muscle, 
especially  in  the  deeper  layers.  From  its  under  surface  a  number  of  trabeculze  carrying  blood- 
vessels pass  into  the  substance  of  the  gland,  thus  dividing  it  into  a  number  of  compartments 
or  alveoli,  which  are  filled  with  a  network  of  delicate  adenoid  tissue,  whose  meshes  are  in 
great  part  filled  with  lymph-corpuscles.  The  gland-substance  consists  of  a  cortical  and  a 
medullary  part.  In  the  cortex  the  adenoid  tissue  is  arranged  in  the  form  of  small,  round,  or 
oval  nodules  ox  follicles,  and  in  the  medulla  as  interlacing  cords.  The  lymph  passes  through 
well-marked  channels  in  the  gland — the  lymph-spaces — which  are  spaces  lying  between  the 
septa  or  trabeculse  and  the  gland-substance.  These  spaces  are  lined  by  squamous  epithelium, 
and  across  them  is  stretched  adenoid  tissue,  which  contains  few  lymph-corpuscles.  The 
structure  of  adenoid  tissue  has  already  been  considered  (p.  29). 

PREPARATION  {a). — Place  a  lymphatic  gland  from  an  o.x  or  other  animal  in  Muller's 
fluid  for  a  week,  and  then  harden  it  in  spirit  and  make  sections.  (F)  Harden  a  gland  in  a 
saturated  watery  solution  of  picric  acid  for  twenty-four  hours,  and  preserve  in  alcohol.  Stain 
a  Mijller's  fluid  section  with  log\vood  and  mount  it  in  dammar,  and  stain  a  picric  acid  section 
with  picrocarmine  and  mount  it  in  Farrant's  solution. 

Lymphatic  Gland.  (Logwood  and  dammar.)  EXAMINATION  (L). — Observe  the  capsule 
with  its  septa  subdividing  the  corte.x  into  a  number  of  compartments  o{  alveoli.  As  they  pass 
into  the  centre  of  the  gland  the  septa  break  up  into  trabeculcB,  which  run  in  all  directions,  and 
anastomose  with  each'  other,  forming  a  plexus  with  relatively  small  meshes,  so  that  it  is  easy  to 
distinguish  the  small-meshed  medulla  from  the  cortex.  Observe  the  larger  alveoli  in  the 
cortex.  Notice  the  masses  of  leucocytes  stained  deep  blue,  and  between  these  and  the 
trabeculae  narrow  spaces,  the  lymph-sinuses.  {Indicate  the  capsule,  cortex,  and  medulla  iri  PI. 
XVII.,  Fig.  3).  (H).  Obser\-e  the  two  layers  of  the  capsule;  the  superficial  layer  contains 
connective  tissue  and  a  few  elastic  fibres,  the  deep  one  also  non-striped  muscular  fibres.  Study 
one  of  the  trabecular,  which  consist  of  connective  tissue  with  a  few  elastic  fibres  ;  and  observe 
the  nuclei  of  the  non-striped  muscle.     The  trabecute  arc  cut  in  every  direction,  and  branch 


78  PRACTICAL   HISTOLOGY. 

frequently.  Continuous  on  each  side  with  the  trabeculse  is  the  adenoid  tissue,  stretching 
across  the  lymph-space  (PI.  XVII.,  Fig.  4).  Blue-stained  nuclei  are  seen  at  the  nodal  points 
of  the  fibres  (PI.  XVII.,  Fig.  6).  As  already  shown  (p.  29),  they  are  the  nuclei  of  squames 
which  cover  these  fibres.  The  me.shes  contain  few  leucocytes.  Study  a  patch  loaded  with 
lymph-corpuscles,  and  also  a  transverse  section  of  a  trabecula  with  its  lymph-space  around 
it  (PI.  XVII.,  Fig.  5).     This  is  most  easily  found  in  the  medulla. 

The  picric  acid  and  picrocarmine  section  shows  similar  details. 

Refer  also  to  the  preparations  of  adenoid  tissue  prepared  by  the  interstitial  injection  of  a 
solution  of  silver  nitrate,  which  demonstrates  the  endothelium  lining  the  lymph-sinuses  (p.  29). 

Injection  of  the  Lymph-Sinuses.  PREPARATION. — With  a  hypodermic  syringe  inject  into 
the  lymphatic  gland  of  an  ox  a  two  per  cent,  watery  solution  of  Prussian  blue.  Place  the 
gland  in  alcohol  for  twenty-four  hours,  and  then  make  transverse  sections.  Stain  a  section 
with  carmine,  and  mount  it  in  Farrant's  solution. 

EXAMINATION  (L). — Observe  the  distribution  of  the  blue  deposit  around  the  follicles  in 
the  cortex,  and  around  the  trabeculae  in  the  medulla.  This  method  indicates  beautifully  the 
position  of  the  lymph-sinuses. 

Blood-vessels. — Study  a  section  where  the  blood-vessels  have  been  injected,  and  note  that 
the  large  vessels  lie  in  the  trabeculae,  whilst  the  capillaries  are  almost  entirely  confined  to  the 
glandular  substance. 

(B.)     SIMPLE    LYMPHATIC    GLANDS. 

These  consist  of  masses  of  adenoid  tissue  occurring  in  the  form  of  spherical  or  nodular 
masses,  either  singly  or  in  groups,  or  sometimes  in  the  form  of  cords.  They  have  already 
been  referred  to  as  occurring  in  the  tonsils  (p.  65),  at  the  root  of  the  tongue  (p.  61),  epiglottis 
(P-  S3),  trachea  (p.  54),  and  lungs  (p.  59),  (especially  in  the  lungs  of  the  cat),  in  the  pyloric 
end  of  the  stomach  (p.  6j),  throughout  the  intestine  as  solitary  glands  (p.  70),  and  in  groups, 
as  Peyer's  patches,  in  the  small  intestine  (p.  69).  They  also  occur  in  the  spleen  (p.  yg)  as 
Malpighian  corpuscles.     They  form  the  basis  of  the  thymus  gland  (p.  81). 


THE    SPLEEN. 

It  is  desirable  to  examine  the  spleen  of  one  of  the  lower  animals,  say  a  cat,  as  well  as  that 
of  man. 

(A.)    CAT. 

PREPARATION. —  Cut  the  spleen  of  a  cat  into  five  or  six  pieces,  and  harden  them  first  in 
Miiller's  fluid  ;  at  the  end  of  a  week  place  them  in  chromic  acid  and  spirit  mixture,  and  after 
another  week  transfer  them  to  spirit.  Make  transverse  sections.  Stain  a  section  in  logwood 
and  mount  it  in  dammar. 

(B.)     HUMAN. 

Cut  a  fresh  normal  human  spleen  into  small  pieces,  and  harden  them  in  the  same  way  as 
the  above,  but  leave  them  two  weeks  in  the  chromic  acid  and  spirit  fluid.  Make  sections  of  a 
part,  and  include  the  fibrous  capsule.  Stain  a  section  in  logwood,  and  mount  it  in  Farrant's 
solution. 


THE  SPLEEN.  79 

Transverse  Section  of  a  Cat's  Spleen.  EXAMINATION  (L). — Observe  the  serous  covering 
and  also  the  fibrous  capsule,  thick  and  firmly  adherent  to  the  subjacent  organ.  Trace  the 
large  coarse  fibrous  traheculce  passing  from  its  under  surface  into  the  organ,  where  they  anas- 
tomose and  form  a  trabecular  framework,  which  divides  the  organ  into  compartments  of  unequal 
size,  that  communicate  with  each  other,  and  are  filled  with  the  splenic  pulp.  The  splenic 
pulp  consists  of  a  large  number  of  cells  like  lymph-corpuscles,  mixed  with  yellow  blood- 
corpuscles.  In  it  observe  the  rounded  aggregations  of  lymph-corpuscles  constituting  Mal- 
pighian  or  splenic  bodies.  They  are  far  more  numerous  relatively  than  in  the  human  spleen, 
and  lie  scattered  irregularly  in  the  pulp.  They  are  not,  as  occasionally  described,  always 
spherical  bodies,  but  in  reality  are  cord-like  masses  of  adenoid  tissue,  like  those  of  the  lung, 
developed  on  the  walls  of  the  arteries,  usually  more  on  one  side  of  it  than  on  the  other,  so  that 
one  must  look  for  a  section  of  an  artery  in  each  mass  ;  and  the  position  of  the  artery  is  eccentric. 
Sometimes  they  are  oval  or  spherical  in  shape.  In  the  trabeculae  search  for  sections  of  the 
large  branches  of  the  splenic  artery  and  vein,  which  run  for  a  certain  distance  together  in  these 
trabeculae.  It  is  to  be  observed  that  the  connective  tissue  which  passes  in  on  the  blood-vessels 
at  the  hilum  becomes  continuous  with  the  trabecule  of  the  spleen  itself.  There  is  no  lymph- 
space  between  the  trabecular  and  the  pulp.  (^Indicate  the  general  arrangement  iji  one  half  of 
PI.  XVIII.,  Fig.  I.) 

(H).  Observe  the  capsule,  consisting  of  several  layers  of  fibrous  tissue  intermixed  with  a 
few  elastic  fibres  and  some  non-striped  muscle.  Select  a  Malpighian  body.  Observe  that  it 
is  made  up  of  leucocytes,  lying  in  a  meshwork  of  adenoid  tissue.  It  has  no  definite  wall,  its 
outer  boundary  is  indicated  by  the  leucocytes  being  more  crowded  together,  and  so  it  is 
stained  darker.  It  is  a  mass  of  adenoid  tissue  developed  in  the  adventitia  of  an  artery  ; 
therefore  look  for  a  section  of  an  artery  in  it,  cut  either  transversely  or  longitudinally. — The 
Ptdp.  Observe  the  colourless  corpuscles,  and  note  the  admixture  of  a  large  number  of 
blood-corpuscles.  The  fine  network  of  delicate  fibrils  in  which  they  lie  imbedded  may  be 
observed. 

Human  Spleen.  (Logwood  and  Farrant's  solution.)  EXAMINATION  (L). — Observe  the 
capsule,  trabeculae,  and  splenic  pulp.  In  the  latter  notice  a  mottled  yellow  and  bluish  ap- 
pearance. The  yellow  streaks  indicate  the  position  of  the  blood-stream  in  the  splenic  pulp. 
All  the  leucocytes  are  stained  blue.  Observe  the  relatively  small  number  of  splenic  corpuscles, 
which  are  usually  ill-defined.  The  splenic  corpuscles  can  usually  be  best  seen  in  the  spleen 
of  a  young  person,  especially  if  the  spleen  be  congested  ;  small  masses  of  yellowish-brown 
pigment  may  be  found  in  the  pulp. 

(H).  A  Malpighian  corpuscle  consists  of  a  vascular  mass  of  adenoid  tissue,  viz.,  a  mesh- 
work of  adenoid  reticulum  loaded  with  leucocytes,  many  of  which  have  two  spherical  nuclei 
(PI.  XVIII.,  Fig.  3).  At  their  periphery  they  shade  into  the  splenic  pulp.  The  splenic  pulp  is 
the  most  difficult  part  of  the  spleen  to  understand.  Select  a  thin  fragment  at  the  margin  of 
the  section,  and  observe  the  fine  network  of  delicate  fibrils  crowded  with  leucocytes,  and  a 
very  large  number  of  coloured  blood-corpuscles.  The  latter  give  the  pulp  its  yellow  mottled 
appearance.  The  framework  or  matrix  of  the  pulp,  when  seen  in  sections,  appears  to  consist  of 
fine  fibrils,  which  form  a  network  with  very  small  meshes,  varying  in  size  from  a  coloured 
blood-corpuscle  to  three  or  four  times  that  size.  It  is  probable,  however,  that  it  consists  of  nu- 
cleated, branched,  cell-plates,  which  anastomose  so  as  to  produce  a  honeycomb  arrangement 
(Klein).  They  not  unfrequently  contain  (especially  in  the  congested  human  spleen)  small 
masses  of  fine  or  coarse  yellow  pigment.  It  is  important  to  observe  that  this  fine  meshwork 
is  directly  continuous  with  the  radicles  of  the  veinlets.     The  cell-plates  arrange  themselves  into 


8o  PRACTICAL    HISTOLOGY. 

the  form  of  a  continuous  tube,  and  at  the  same  time  they  become  more  spindle-shaped.  This 
is  the  mode  of  origin  of  a  venous  radicle  from  the  pulp,  so  that  the  spaces  in  the  splenic  pulp 
are  directly  continuous  with  the  vascular  system.    {Indicate  these  details  in  PI.  XVIII.,  Fig.  2.) 

BLOOD-VESSELS    OF   THE    SPLEEN. 

The,  splenic  artery  enters  the  organ  (at  the  hilum)  along  with  the  vein,  lymphatics,  and 
nerves,  all  of  which  are  imbedded  in  the  connective  tissue,  which  becomes  continuous  with  the 
splenic  trabeculae.  It  runs  for  a  short  distance  in  these  trabeculas,  but  it  soon  leaves  the  vein, 
and  its  branches  divide  suddenly  into  a  large  number  of  smaller  branches  (pcnicilli).  The  small 
branches  soon  become  more  or  less  completely  ensheathed  by  a  solid  mass  of  adenoid  tissue, 
and  this  tissue  constitutes  the  so-called  Malpighian  or  splenic  corpuscles.  These  are  supplied 
by  distinct  capillaries,  which  form  a  uniform  network  with  wide  meshes.  Other  arterial 
branches  open  directly  into  the  meshwork,  or  '  honeycombed  matrix,'  of  the  splenic  pulp, 
which,  as  above  described,  opens  into  the  rootlets  of  the  veins  (venous  sinuses),  and  these 
again  into  the  larger  venous  trunks  lying  in  the  trabeculae.  No  capillaries  are  found  in  the 
pulp  ;  they  are  confined  to  the  Malpighian  corpuscles.  At  the  periphery  of  these  corpuscles 
the  capillaries  open  directly  into  the  spaces  of  the  pulp,  which  are  therefore  the  channels  of 
communication  between  the  terminations  of  the  arteries  and  the  origin  of  the  venous  radicles. 
The  blood-stream  in  the  splenic  pulp,  therefore,  corresponds  to  the  lymph-stream  in  a 
lymphatic  gland. 

PREPARATION.  Injection  of  the  Blood-vessels  of  the  Spleen,  (a)  Carmine  Gelatine  Mass. — 
It  is  well  to  use  a  constant-pressure  apparatus,  and  to  cease  injecting  whenever  the  mass 
ceases  to  flow  into  the  spleen.  Inject  from  the  splenic  artery  a  thin  carmine  gelatine  mass. 
If  the  human  spleen  be  taken,  select  the  spleen  of  a  child.  After  the  injection  is  completed 
harden  the  organ  in  Mijller's  fluid.  The  spleen  of  a  rat  does  very  well,  but  in  all  cases  the 
animal  ought  to  be  killed  by  bleeding.  In  the  case  of  animals  it  is  sometimes  advantageous 
to  wash  out  the  blood-corpuscles  of  the  spleen  with  a  stream  of  salt  solution  before  using  the 
gelatine  mass. 

ip)  Silver  Nitrate  Injection. — Wash  out  the  blood-vessels  with  distilled  water,  and  then 
throw  in  a  quarter  per  cent,  solution  of  silver  nitrate.  Harden  in  alcohol.  Sections  of  this 
show  the  endothelium  of  the  venous  sinuses. 

Make  sections  of  the  above,  stain  them  with  logwood,  and  mount  them  in  dammar. 

EXAMINATION  (L). — Observe  the  Malpighian  corpuscles  ;  oval  or  irregularly  shaped 
bodies  stained  blue,  and  in  each  note  a  branch  of  the  splenic  artery  and  a  few  capillaries. 
The  pulp  contains  a  network  of  spaces  (venous  sinuses),  filled  with  the  red  carmine  mass,  and 
in  this  lie  the  masses  of  logwood-coloured  '  pulp  '  of  the  spleen,  which  are  about  the  same 
breadth  as  the  venous  or  cavernous  sinuses.  Larger  spaces — the  rootlets  of  a  vein — may 
be  seen  opening  directly  into  these  spaces.  {Indicate  the  blood-vessels  in  one  half  of  PI.  XVIII., 
Fig.  I.) 

(H).  Note  the  network  of  spaces  filled  with  the  red  mass,  and  between  them  the  lymph- 
cells  stained  blue.  Select  a  venous  radicle,  and  note  the  layer  of  endothelium  lining  it.  Trace 
the  continuation  of  an  intra-Malpighian  capillary  into  the  red  network  at  its  margin. 

NERVES    OF    THE    SPLEEN. 

These  have  already  been  alluded  to  (p.  45).  Amongst  the  non-medullated  nerve-fibres,  I 
have  found  a  large  number  of '  plasma-cells '  (p.  20). 


PLATE  X^/ffl    SPHEEN  &^ii-ij.r^-aE  GlAED^ 


Mintem.  Bros  lith. 


THE  SPLEEN.  8i 


FRESH    SPLEEN. 

Cut  across  a  fresh  ox  spleen.  Observe  the  capsule,  trabeculae,  and  dark  grumous-like  pulp. 
The  pulp  can  easily  be  washed  away  with  a  stream  of  water  or  salt-solution  from  a  wash-bottle  ; 
the  trabecular  framework  is  then  easily  observed.  Observe  the  Malpighian  corpuscles, 
small  white  bodies  about  the  size  of  the  head  of  a  pin.  Do  not  mistake  transverse  sections  of 
the  trabeculae  for  them.  Pick  out  one,  and  examine  it  in  salt  solution  (H).  It  is  seen  to  be 
made  up  of  lymph-corpuscles,  as  already  described. 

SPLENIC    PULP. 

Tease  a  small  part  of  fresh  splenic  pulp  in  salt  solution,  and  apply  a  cover-glass. 

EXAMINATION  (H). — Observe  the  lymph-corpuscles,  perhaps  the  endothelial  cells  from  a 
vein,  also  cells  containing  particles  of  fat,  others  with  coloured  blood-corpuscles,  and  some 
containing  pigment,  besides  numerous  coloured  blood-corpuscles.  If  the  coloured  blood- 
corpuscles  are  so  numerous  as  to  obscure  the  view  of  the  other  elements,  tease  the  pulp  in 
water,  which  dissolves  the  blood-corpuscles  ;  thus  the  other  elements  can  then  be  seen  more 
distinctly. 


THE    THYMUS. 

The  thymus  consists  of  an  aggregation  of  lymph-follicles,  or  masses  of  adenoid  tissue 
held  together  by  connective  tissue,  which  contains  blood-vessels,  lymphatics,  and  a  few  nerves 
The  framework  consists  of  a  capsule  of  fibrous  tissue,  which  gives  off  septa  dividing  the 
gland  into  lobes,  these  being  further  subdivided  by  finer  septa  into  lobules,  the  lobules  being 
subdivided  by  fine  intra-lobular  lamellse  of  connective  tissue  into  follicles.  The  gland- 
substance  is  made  up  oi  \hcsQ  follicles,  which  are  more  or  \es,s  polygonal  in  outline  from  mutual 
pressure.  Each  follicle  consists  of  a  cortical  and  a  medullary  part,  and  the  matrix  or  frame- 
work of  each  consists  o{  2.  fine  adenoid  reticitliiin  (p.  29),  made  up  of  nucleated  branched  cells, 
which  are  easily  seen.  In  this  meshwork  the  lymph-corpuscles  lie.  In  the  medulla  are  found 
the  so-called  concentric  corpjiscles  (Hassall).  They  consist  of  a  central  granular  part,  around 
which  are  disposed  concentrically  layers  of  flattened  nucleated  endothelial  cells,  and  on  section 
they  present  appearances  like  the  '  cell-nests '  that  occur  in  epithelioma. 

PREPARATION  {a). — Harden  part  of  the  thymus  of  an  infant  in  chromic  acid  and  spirit 
mixture  for  two  weeks,  and  then  transfer  it  to  alcohol.  Stain  a  section  with  logwood  and 
mount  it  in  dammar. 

[li)  Harden  a  similar  piece  in  picr'c  acid  for  twenty-four  hours,  and  then  transfer  it  to 
spirit.  Sections  of  this  may  be  beautifully  stained  with  picrocarmine.  Mount  it  in  Farrant's 
solution.  The  thymus  of  a  kitten  or  puppy  may  be  used,  but  these  do  not  show  the  concentric 
corpuscles  so  well. 

EXAMINATION  (L).— Observe  the  capsule,  the  interlobular  and  intralobular  septa  of 
connective  tissue,  made  up  of  lamellae  with  flattened  corpuscles  between  them.  Note  the 
polygonal  or  irregularly  shaped  follicles.  Study  a  follicle.  Observe  the  cortical  part  stained 
more  deeply  with   the  dye   than   the   medulla.     (H).   Search   for  the    adenoid    reticulum    of 

M 


82  PRACTICAL   HISTOLOGY. 

branched  cells,  and  study  the  concentric  corpuscles.     The  reticulum  is  best  seen  in  prepara- 
tions mounted  in  Farrant's  solution  or  glycerine. 

The  concentric  corpuscles  can  easily  be  isolated  by  teasing  a  small  part  of  a  fresh  gland 
in  salt  solution. 


THE    DUCTLESS   GLANDS. 

These  are  the  thyroid  gland,  hypophysis  cerebri,  \}n<t  coccyge a/  and  carotid  glands  of  Luschka, 
and  the  supra-renal  body.  They  have  no  special  ducts,  and  their  secretions  are  probably  re- 
moved by  the  lymphatics. 

THE    THYROID    GLAND. 

It  consists  of  a  capsule  and  a  connective-tissue  framework,  which  subdivides  the  glands 
into  lobes  and  lobules.  Each  lobule  is  made  up  of  a  larger  or  smaller  number  of  gland-alveoli 
or  gland-vesicles,  which  are  closed  sacs  of  various  sizes,  and  are  spherical,  oval,  or  irregular  in 
shape.  Each  sac  or  alveolus  consists  of  a  membrana  propria  lined  by  a  single  layer  of  more 
or  less  columnar  nucleated  cells.  The  size  of  the  central  lumen  depends  essentially  on  the 
amount  of  secretion.  These  sacs  are  filled  with  a  transparent,  albuminous,  viscid,  slightly 
yellow-coloured  fluid.  It  is  the  accumulation  of  this  fluid,  and  the  consequent  distension  of 
these  sacs,  which  constitutes  goitre.  Not  unfrequently  the  alveoli  contain  large  numbers  of 
coloured  blood-corpuscles  (Baber).  Each  alveolus  is  surrounded  by  a  rich  plexus  of  capillaries 
which  do  not  penetrate  the  membrana  propria.  As  in  other  glands,  the  large  blood-vessels, 
lymphatics,  and  nerves  lie  in  the  interlobular  connective  tissue. 

PREPARATION.— Harden  the  thyroid  of  a  child  in  Muller's  fluid  for  three  weeks,  or  in 
chromic  acid  and  spirit  mixture  for  two  weeks.  Then  transfer  it  to  spirit.  Make  sections, 
stain  them  with  logwood,  and  mount  them  in  dammar. 

EXAMINATION  (L). — Observe  the  interlobular  connective  tissue  and  the  gland-vesicles, 
which  are  of  difierent  sizes  and  lined  by  a  single  layer  of  columnar  epithelium.  (H). 
Study  the  membrana  propria  and  epithelium  of  the  gland-vesicles  (PI.  XVIII.,  Fig.  4). 

Blood-vessels  of  the  Thyroid. — They  are  easily  injected  when  the  whole  body  of  a  child  or 
young  animal  is  injected,  but  the  blood-vessels  are  usually  so  filled  naturally  with  blood  that 
an  injection  need  not  be  made.     Mount  sections — stained  or  unstained — in  dammar. 


THE   SUPRA-RENAL   CAPSULES. 

The  frameivork  consists  of  a  fibrous  tissue  capsule,  which  sends  trabeculae  into  the 
interior  of  the  organ,  which  is  divided  into  a  cortex  and  a  medulla.  In  the  outermost  zone  of 
the  cortex  the  trabeculae  form  a  plexus  with  polygonal  meshes,  whilst  in  the  middle  zone 
they  are  thinner  and  have  a  longitudinal  arrangement,  i.e.  radiate  inwards  ;  in  the  innermost 
zone  of  the  cortex  they  again  form  a  polygonal  meshwork.  A  similar  plexus  of  trabeculae 
exists  in  the  medulla.  These  meshes  are  filled  with  the  parencliyuia  or  gland-substance,  which 
varies  in  appearance  in  the  different  zones.  In  the  outer  cortical  zone  they  are  nucleated, 
granular,  polyhedral  cells  ;  in  the  middle  zone  transparent  nucleated  cells,  often  containing 
oil-globules,  and  arranged  in  the  form  of  cylinders,  which  anastomose  with  each  other  ;  whilst 
in  the  inner  cortical  zone  the  cells  frequently  contain  a  yellowish-brown  pigment.     With  these 


SUPRA-RENAL    CAPSULES.  8 


o 


are  continuous  the  transparent,  brittle,  and  sometimes  pigmented  and  branched  cells  of  the 
medulla.  Both  cortex  and  medulla  are  richly  supplied  with  blood-vessels,  the  distribution 
of  which  follows  that  of  the  trabeculae.  A  large  number  of  nerves  and  nerve-ganglia  exists 
in  the  organs. 

PREPARATION.—  It  is  best  to  emploj-  the  capsules  of  large  mammals,  e.jf.  of  the  ox,  or, 
best  of  all,  the  horse.  If  these  cannot  be  obtained,  use  the  supra-renals  of  a  guinea-pig.  In 
the  case  of  the  human  organ,  it  must  be  obtained  as  fresh  as  possible,  (a)  Harden  pieces  in 
Miiller's  fluid  for  three  weeks,  or  in  chromic  acid  and  spirit  mixture  for  two  weeks,  and  then 
transfer  them  to  spirit.  Make  transverse  sections,  and  stain  them  with  logwood,  and  mount 
them  in  dammar. 

(d)  Osmic  Acid. —  Place  pieces  about  the  size  of  a  pea  in  a  quarter  per  cent,  osmic  acid  for 
eight  hours,  and  complete  the  hardening  in  spirit.  This  method  gives  excellent  results,  but  it 
must  be  remembered  that  the  osmic  acid  is  apt  to  fix  only  the  outer  layers. 

EXAMINATION  (L).— Observe  the  capsule  and  its  trabecule;,  also  the  cortex  with  its 
three  zones  and  the  medulla,  and  note  the  partially  pigrnented  cells  (PI.  XX.,  Fig.  4).  (H). 
Study  the  cylinders  of  cells,  and  compare  them  with  the  foregoing  description  (PI.  XX., 
Fig.  5). 

Study  the  injected  specimen,  and  note  the  distribution  of  the  blood-vessels. 


84  PRACTICAL   HISTOLOGY. 


THE   KIDNEYS. 

THE    COURSE    AND    THE    STRUCTURE    OF    THE    URINARY 

TUBULES. 

The  course  of  the  urinifeious  tubules  is  very  complicated,  and  the  epithelium  differs  in  different 
sections  of  the  tubules. 

The  urinary  tubules  begin  as  a  blind  extremity  at  the  Malpighian  corpuscle,  and  open  on 
the  free  surface  of  a  papilla.  Every  tubule  consists  of  the  following  sections,  which  are  con- 
tinuous with  each  other. 

(i)  The  Malpighian  capsule,  \v\\\c\\  passes  by  (2)  a  short,  narrow  neck  or  constrictio-n  into  (3) 
the  proximal  convoluted  tube,  which  passes  into  (4)  the  spiral  tubule  (Schachowa),  lying  in 
a  pyramid  of  Ferrein.  All  the  above  lie  in  the  cortex.  As  the  spiral  tube  passes  from  the 
cortex  into  the  boundary  layer,  it  suddenly  becomes  straight  and  narrow  as  (5)  the  descending 
limb  of  Henles  loop,  which  is  continued  into  the  beginning  of  the  papillary  portion,  where  it 
bends  on  itself  and  forms  (6)  Henles  loop,  which  ascends  into  the  boundary  layer,  where  it 
suddenly  enlarges  and  becomes  slightly  wavy,  to  form  (7)  the  first  thick pa7-t  of  t lie  ascending 
limb  of  Henles  loop.  It  again  becomes  narrow  and  spiral,  when  it  forms  (8)  the  spiral  part  of 
the  ascending  limb.  It  now  (9)  re-enters  the  cortex,  and,  becoming  narrower,  ascends  in  a 
pyramid  of  Ferrein,  which  it  soon  leaves,  and  enters  the  labyrinth  amongst  the  convoluted 
tubules,  as  (10)  the  irregular  tube,  so  called  because  of  its  irregular  size,  breadth,  and  course. 
This  passes  into  (\\)  the  intercalated  segment  (Schweigger-Seidel)  which  is  the  distal  convoluted 
tube,  exactly  like  (3).  It  passes  into  (12)  the  curved  part  of  the  collecting  tube,  a  narrow  curved 
tube  which  passes  through  the  labyrinth,  where  it  is  joined  by  other  similar  tubes,  and  forms 
(13)  the  straight  part  of  the  collecting  tube,  which  descends  in  a  pyramid  of  Ferrein,  and  then 
enters  the  boundary  portion,  and  finally  passes  into  the  papillary  portion,  there  to  be  joined  by 
other  branches,  and  form  a  large  (14)  collecting  tube,  opening  on  the  free  surface  of  a  papilla 
(Klein). 

The  membrana  propria  of  Bowman's  capsule  is  continuous  throughout  all  the  sections  of 
the  urinary  tubules,  but  its  epithelial  lining  varies  greatly  in  different  parts  of  its  course. 

Bowman's  capsule  is  lined  with  a  single  layer  of  squames  ;  the  proximal  convoluted  tubule 
(3)  with  a  single  layer  of  low  nucleated  columnar  epithelial  cells,  which  leave  a  lumen  the  size 
of  one-third  the  diameter  of  the  tube.  All  the  other  sections  of  the  tube,  except  the  de- 
scending part  of  Henle's  loop  (s),  and  all  the  parts  of  the  collecting  tube  (12,  13,  and  14)  are  lined 
with  polyhedral  nucleated  epithelial  cells,  whose  outer  half  contains  vertically  placed  '  rods '  or 
fibrils  (p.  87). 

The  descending  limb  of  Henle's  loop  (5  and  6)  is  lined  with  a  layer  of  very  flat  transparent 


KIDNEYS.  85 

nucleated  plates,  and  therefore  not  unlike  blood-vessels.  The  ascending  limb  (7)  in  its 
wide  part  is  lined  with  polyhedral  '  rodded  '  cells,  with  their  nuclei  next  the  innermost  part  of 
the  cells.  The  narrow  ascending  part  (8)  is  lined  with  low  polyhedral  cells,  and  has  a  ver>' 
narrow  lumen.  The  collecting  tubes  are  lined  with  a  single  layer  of  homogeneous  transparent 
nucleated  cells,  which  vary  considerably  in  shape. 

It  is  to  be  remembered  that  the  above  arrangement  has  only  been  made  out  after  laborious 
investigations,  and  no  such  complete  tubule  was  ever  seen,  or  can  ever  be  seen,  in  a  section  of 
a  kidne)'.  The  course  of  the  tubules  can  only  be  made  out  by  isolating  the  tubules  as 
described  at  p.  86. 

PREPARATION,  a.  Chromic  Acid  and  Spirit  Mixture. — Remove  the  kidneys  from  an  animal 
just  killed  by  bleeding — rabbit,  dog,  or  cat  ;  divide  one  transversely  into  several  pieces  with  a 
sharp  razor,  and  place  them  in  the  above  fluid.  Take  great  care,  in  order  not  to  separate  the 
capsule.  Divide  the  other  longitudinally  and  treat  it  in  the  same  way.  Make  both  transverse 
and  longitudinal  sections  of  the  kidney.  Stain  some  with  logwood  and  mount  either  in 
dammar  or  in  Farrant's  solution,  whilst  others  may  be  stained  with  picrocarmine  and  mounted 
in  Farrant's  solution. 

b.  Miiller's  fluid  may  be  employed  in  the  same  way  as  the  above,  but  takes  from  three  to 
four  weeks  to  complete  the  hardening. 

c.  Ammonium  Chromate  (Heidenhain).  Harden  small  pieces  in  a  five  per  cent,  solution  of 
ammonium  chromate  for  forty-eight  hours,  then  wash  away  all  the  colouring  matter,  and 
complete  the  hardening  in  dilute,  and  then  in  strong  spirit. 

Prepare  portions  of  a  normal  hiniian  kidney  in  the  same  way  {a  and  c).  Compare  sections 
of  these  with  the  above. 

KIDNEY    OF   A   RABBIT    OR    DOG. 

Transverse  Section.  EXAMINATION.— With  the  naked  eye  note  the  general  shape  of  the 
section,  distinguishing  the  capsule,  which  is  very  easily  detached  from  the  cortex.  Note  the 
division  of  the  pare?ichyiiia  into  three  distinct  regions  :  the  cortex,  the  boundary  layer  (Ludwig) 
and  the  papillary  portion,  the  two  last  forming  the  medulla,  which  terminates  in  a  papilla, 
projecting  into  the  pelvis  of  the  kidney. 

(L).  In  the  medulla  observe  the  radiating  rows  of  straight  tubules  (tubuli  recti)  proceeding 
from  the  pelvis  outwards.  As  they  pass  outwards  they  divide  dichotomously.  When  taken 
collectively  they  constitute  a  Malpighian  pyramid..  In  the  r<7;Yf.r  observe  that  on  the  boundary 
line  between  it  and  the  medulla,  the  straight  tubules  of  the  Malpighian  pyramid  are  continued 
as  bundles  of  straight  tubules  into  the  cortex,  constituting  the  pyramids  of  Ferrein,  or  me- 
didlary  rays,  which  are  the  direct  continuation  of  the  straight  tubes  of  the  medulla  into  the 
cortex.  As  they  radiate  towards  the  capsule  these  bundles  of  tubules  become  smaller,  and 
are  always  separated  from  the  capsule  by  a  layer  of  convoluted  tubules.  Observe  the  convo- 
luted tubules  [tubuli  contorti)  confined  to  the  cortex.  They  lie  between  the  pyramids  of 
Ferrein,  and  between  their  upper  ends  and  the  capsule.  Between  two  pyramids  of  Ferrein 
observe  the  double  row  of  Malpighian  corpuscles,  which  consist  of  the  capsule  of  Boiuman  in- 
closing a  glomerulus,  or  tuft  of  blood-vessels.  Sometimes  the  glomeruli  fall  out,  and  then 
there  is  only  a  circular  aperture  in  the  section.  Notice  under  the  capsule  more  or  less  tri- 
angular spaces,  which  are  transverse  sections  of  veins.  Observe  how  easily  the  capsule  sepa- 
rates, and  note  the  very  limited  amount  of  connective  tissue  which  passes  from  its  under 
surface  into  the  organ.  {Indicate  the  shape  of  the  kidney  and  the  general  arrangement  of  the 
tubules  in  PI.  XIX.,  Fig.  i.) 


86  PRACTICAL    HISTOLOGY. 


HUMAN    KIDNEY. 

Examine  (L)a  similar  section  of  a  human  kidney,  and  note  especially  the  normal  thickness 
of  the  capsule  and  its  delicate  and  scanty  fibres  of  attachment  to  the  subjacent  organ.  {Indicate 
the  capsule,  pyramids  of  Ferrein,  convoluted  tubules,  and  Malpighian  corpuscles  in  PI.  XIX., 
Fig.  2.) 

(H).  Select  a  Malpighian  corpuscle.  Observe  the  membrane  or  capsule  of  Bowman,  con- 
sisting of  an  elastic  membrana  propria,  and  note  the  single  layer  of  squamous  epithelium 
lining  it.  The  nuclei  are  stained,  and  bulge  slightly  into  the  cavity.  Outside  it  there  is  a 
small  amount  of  fibrous  tissue.  Within  it  note  the  tuft  of  blood-vessels,  with  their  nuclei 
stained,  and  the  capillaries,  perhaps,  containing  yellow  blood-corpuscles  (PI.  XIX.,  Fig.  3). 
The  capillaries  are  arranged  in  two  or  three  conical  lobules,  but  do  not  completely  fill  the 
capsule  ;  the  space  between  the  surface  of  the  glomerulus  and  the  capsule  depends  on  the 
amount  and  nature  of  the  secretion  present  in  it.  Between  the  capillaries  there  is  a  ver\'  small 
amount  of  connective  tissue  (chiefly  branched  connective-tissue  cells),  which  holds  them  to- 
gether ;  and  over  them  there  is  a  layer  ofsquames.  The  section  may  pass  through  the  capsule 
in  the  plane  of  the  afferent  or  efferent  blood-vessels  ;  or  the  narrow  neck  which  connects  the 
cavity  of  the  capsule  with  a  convoluted  tubule  may  be  found. 

Select  a  convoluted  tubule  (H). — Observe  them,  one  divided  longitudinally  and  another 
transversely,  and  observe  the  limiting  membrane,  and  how  closely  they  are  packed  in  the 
cortex,  with  very  little  supporting  tissue  between  them.  Study  the  secretory  epithelium  lining 
them,  and  note  that  the  lumen  of  the  tube  is  small,  that  the  epithelial  cells  are  ill-defined  and 
have  no  cell-wall.  They  contain  a  spherical  nucleus  placed  near  the  attached  end  of  the  cell, 
and  their  protoplasm  is  always  cloudy  and  granular.  The  outer  part  of  the  protoplasm  is 
often  striated  ;  this  is  especially  the  case  if  the  section  has  been  hardened  in  ammonium 
chromate.  Some  tubules  may  be  found  with  an  angular  zigzag  bend  in  them  ;  these  are  the 
connecting  tubules.     {Indicate  a  convoluted  tubule  in  PI.  XX.,  Fig.  i.) 

Select  a  Pyramid  of  Ferrein  (H) — Observe  the  straight  tubules,  and  amongst  these  the 
looped  tubules  of  Hcnle,  with  their  descending  limb,  which  is  very  narrow  and  lined  by  flattened 
epfthelium,  with  alternately  projecting  nuclei  on  either  side.  They  are  not  unlike  capillary 
blood-vessels,  though  they  have  a  well-marked  basement-membrane.  It  is  difficult  to  distin- 
guish the  ascending  limb,  as  it  is  so  like  a  collecting  tubule.  The  descending  branch  may  be 
traced  downwards  into  the  medulla.     {Indicate  these  tubules  in  PI.  XX.,  Fig.  i.) 

The  Medulla  (H). — Observe  the  straight  tubules — collecting  or  discharging  tubes — which 
open  on  the  ape.x  of  the  Malpighian  pyramid.  Observe  their  columnar  or  cubical  epithelial 
covering.  The  cells  are  well-defined,  and  contain  a  well-marked  spherical  nucleus,  imbedded 
in  clear  transparent  protoplasm.  There  is  a  well-marked  lumen  in  the  centre  of  the  tubule. 
Observe  the  basement-membrane  in  which  the  cells  are  placed,  and  note  the  relatively  large 
amount  of  connective  tissue  in  the  medulla  compared  with  the  cortex  (PI.  XX.,  Fig.  2). 

Place  unstained  sections  of  a  kidney  in  a  quarter  per  cent,  osmic  acid  for  twenty-four 
hours,  and  mount  them  in  Farrant's  solution. 

ISOLATED    RENAL   TUBULES. 

PREPARATION. — The  arrangement  and  relation  of  these  parts  one  to  another  cannot  be 
made  out  in  a  section  ;  to  do  so  the  kidney  must  be  boiled  for  a  long  time  in  a  mixture 
of  alcohol  and  hydrochloric  acid,  which  dissolves  the  connective  tissue  and  liberates  the 
tubules.     A   very   good  method   is  to   place   pieces  of  a  kidney,  about  the   size   of  a  pea,  in 


Plate  xk.iqdney 


MintKrri  Bros  lul-i 


KIDNEYS.  $7 

pure  hydrochloric  acid  for  twelve  hours,  and  then  leave  them  in  water  for  twenty-four  hours. 
Tease  out  a  small  piece  in  osmic  acid  solution  (one  per  cent.),  and  mount  it  in  Farrant's  solu- 
tion. By  teasing  part  of  a  kidney  prepared  in  this  way  it  has  been  shown  that  the  Malpighian 
corpuscle  is  continuous  with  a  convoluted  secreting  tubule,  which  winds  about  in  the  cortex, 
and  then  descends  in  a  pyramid  of  Ferrein  into  the  medulla,  forming  the  descending  part  of 
the  looped  tubule  of  Henle,  and  reascends  as  the  ascending  part  of  the  loop,  and  after  a 
short  course  as  a  connecting  tube,  with  a  zigzag  direction,  becomes  continuous  with  a  collecting 
tube,  and  descends  through  a  pyramid  of  Ferrein,  and  in  its  course  joins  at  an  acute  angle 
with  similar  tubules,  which  is  thus  a  discharging  tube,  opening  on  the  surface  of  a  papilla. 

CONNECTIVE   TISSUE    OF   THE   KIDNEY. 

Compare  this  section  with  the  above.  Make  transverse  sections  through  the  corte.x,  and  also 
through  the  medulla  near  the  apex  of  a  Malpighian  pyramid,  and  stain  them  with  logwood  or 
picrocarmine,  and  mount  in  Farrant's  solution. 

EXAMINATION  (L).— In  the  cortex.  Observe  the  convoluted  tubules  and  glomeruli  ; 
notice  the  small  amount  of  interlobular  connective  tissue,  and  search  for  a  transverse  section 
of  a  group  of  straight  tubules,  i.e.  of  a  pyramid  of  Ferrein.  Study  the  characters  of  these 
parts  (H). 

(L).  Apex  of  a  Malpighian  Pyramid. — Observe  the  transverse  sections  of  the  discharging 
tubules  lined  with  clear,  low,  columnar  epithelium,  and  a  well-defined  lumen.  Notice  the 
large  amount  of  connective  tissue  between  and  supporting  the  tubules,  forming  a  marked 
contrast  to  the  small  amount  in  the  cortex. 

It  is  desirable  to  have  a  transverse  section  through  the  boundary  line  between  the  corte.x 
and  medulla. 

Observe  the  transverse  sections  of  the  collecting  tubules,  and  here  and  there  the  cut  ends 
of  the  ascending  and  descending  looped  tubules  of  Henle  ;  the  narrow  loop  is  not  unlike  a 
capillary  with  its  bulging  oval  nucleus.  Sections  of  capillaries  may  also  be  found,  but  they 
usually  contain  blood-corpuscles.  Note  the  moderate  amount  of  intertubular  connective 
tissue — intermediate  in  amount  between  that  of  the  cortex  and  medulla. 

Ammonium  Chromate  Kidney. — Heidenhain  showed  that  the  epithelial  cells  lining  a 
uriniferous  tubule — excepting  those  of  the  descending  limb  of  Henle's  loop — the  loop  itself, 
and  all  parts  of  the  collecting  tube,  are  made  up  of  a  number  of  '  rods  '  or  fibrils  placed 
vertically  to  the  long  axis  of  the  tube.  These  rods  are  most  distinct  in  the  outer  half  of  the 
cells,  Le.  next  the  membrana  propria.  Each  cell  contains  a  spherical  nucleus  placed  about  the 
middle  of  its  substarfce. 

EXAMINATION  (L).— Select  a  convoluted  tubule  in  the  cortex,  and  use  (H)  to  see  the 
above-described  characters.     They  are  easily  seen  in  a  properly  prepared  kidney. 

BLOOD-VESSELS    OF    THE    KIDNEY. 

PREPARATION. — Make  transverse  and  longitudinal  sections  of  a  kidney  which  has  been 
injected  from  the  renal  artery  with  a  gelatine  and  carmine  or  Berlin-blue  mass.  Mount  in 
dammar. 

EXAMINATION  (L). — Observe  large  branches  of  the  renal  artery  running  outwards  be- 
tween  the    Malpighian    pyramids ;    from    these   the    interlobular  arteries   proceed    vertically 


88  PRACTICAL    HISTOLOGY. 

outwards  between  the  pyramids  of  Fcrrein.  As  the  interlobular  arteries  proceed  outwards, 
they  give  off  on  all  sides  small,  short  trunks — the  afferent  arteries— io  the  Malpi<jhian 
corpuscles,  which  they  enter  and  split  up  into  capillaries  to  form  the  glomerulus  (p.  85). 
Trace  an  efferent  vessel — venous  vessel — cmerginfj  from  the  capsule  at  that  part  where  the 
afferent  vessel  enters  it.  The  efferent  vessel  on  its  exit  splits  up  into  capillaries,  which  form 
a  dense  network  over  and  around  all  the  convoluted  tubules.  From  this  network  the  renal 
vein  proceeds.  Branches  of  it  will  be  found  accompanying  the  interlobular  arteries.  Note  at 
the  innermost  part  of  the  cortex  bunches  of  vessels,  most  of  which  spring  from  the  renal 
artery  and  run  down  in  groups  into  the  medulla — the  vasa  recta  -  where  they  split  up  into  a 
network  with  large  oblong  meshes  surrounding  the  straight  tubules.  The  medulla  is  not 
nearly  so  vascular  as  the  cortex,  but  the  capillaries  of  the  one  are  continuous  with  those  of 
the  other  (PL  XX.,  Fig.  3). 

FRESH    KIDNEY. 

Make  a  longitudinal  section  of  a  perfectly  fresh  sheep's  kidney.  With  the  naked  eye 
observe  the  capsule  and  its  loose  attachment  to  the  parenchyma.  Note  the  division  of  this 
latter  into  three  zones  (p.  85) — the  cortex,  boundary,  ^.nd  papillary  portion.  Study  the  naked- 
eye  characters  of  each  part.  The  papillary  portion  is  uniformly  vertically  striated,  due  to  the 
straight  tubules  and  blood-vessels  running  straight  towards  the  apex  of  the  pyramid.  The 
boundaiy-layer  is  also  striated  longitudinally,  but  is  marked  by  alternate  opaque  and  light 
bluish-coloured  zones  ;  the  former  are  the  continuations  of  the  straight  tubules  towards  the 
cortex,  whilst  the  latter  are  due  to  the  vasa  recta  (p.  88).  The  cortex  is  of  a  light  brown 
colour,  and  is  finely  granular  in  appearance. 

It  is  important  in  a  human  kidney  to  note  the  relative  thickness  of  these  various  parts. 
If  the  vertical  diameter  of  the  cortex  and  medulla  together  be  represented  by  10,  the  relative 
proportions  of  these  three  zones  from  without  inwards  will  be  3"5  :  2'5  :  4  (Klein).  The 
medulla,  therefore,  is  nearly  twice  as  thick  as  the  cortex. 

EXAMINATION  (L). — Scrape  off  a  small  piece  of  the  cortex,  and  tease  it  in  salt  solution 
with  needles.  Observe  the  convoluted  tubules  with  their  cloudy  epithelium,  their  nuclei  being 
scarcely  visible.  Notice  especially  their  basement-membranes,  which  may  be  seen  in  great 
numbers  in  the  field  as  fine  transparent  membranes,  often  with  folds  in  them.  Study  them 
also  with  (H).  Select  a  glomerulus  and  study  it.  Add  dilute  acetic  acid,  and  observe  the 
effect  on  the  epithelium  of  the  convoluted  tubule,  in  rendering  it  clear  and  transparent  and 
revealing  a  nucleus,  and  also  its  effect  on  the  glomerulus  in  rendering  its  capillaries  clear  and 
transparent,  and  bringing  into  view  their  nuclei. 

Scrape  also  the  medulla,  and  examine  it  in  the  same  way  (L  and  H).  Note  the  straight 
tubules  with  their  well-defined,  clear,  epithelial  cells.  Perhaps  isolated  portions  of  the  looped 
tubules  of  Henle  may  be  obtained. 


THE    URETER. 

PREPARATION. — Tie  one  end  of  the  ureter  of  a  dog  or  cat,  and  slightly  distend  it  with  the 
chromic  acid  and  spirit  mixture,  then  tie  the  other  end  ;  leave  it  for  two  days  in  the  mixture  ; 
then  cut  it  into  pieces  an  inch  long,  and  harden  it  for  two  days  longer,  and  complete  the 
hardening  in  spirit.  Make  transverse  sections,  and  stain  some  with  logwood,  and  mount  them 
in  dammar. 


PLATEXX.'KDNEYS^SUPRi^-RENAL   CAPSULE 


Mintcrrt  Bros.lith. 


BLADDER.  89 

EXAMINATION  (L). — Note  the  (i)  external  fibrous  coat  of  connective  tissue;  the  (2) 
middle  or  muscular  coat,  consisting  of  non-striped  muscle  arranged  in  two  layers — («)  an 
external  circular,  and  {J})  an  inner  longitudinal,  with  the  muscle-cells  composing  it  cut 
transversely  ;  and  (3)  the  internal  or  mucous  coat  lined  by  stratified  transitional  epithelium. 
(H).  Study  each  of  these  coats,  but  note  the  transitional  epithelium  (p.  12) ;  the  upper  cells 
are  polyhedral,  and  below  them  is  a  layer  of  pear-shaped,  or  club-shaped,  cells. 


THE    BLADDER. 

PREPARATION. — Prepare  this  exactly  in  the  same  way  as  the  ureter.  Use  the  bladder  of 
a  cat  or  dog.  It  must  be  cut  into  several  pieces  after  being  distended  for  two  days.  Make 
vertical  sections,  and  stain  them  with  logwood,  and  mount  in  Farrant's  solution. 

It  has  practically  the  same  structure  as  the  ureter.  Outside  is  the  ( i )  serous  coat  ;  (2)  the 
7njiscular  coat,  with  its  fibres  arranged  in  three  directions  ;  (3)  the  submucous  coat ;  (4)  the 
mucous  coat  lined  with  stratified  transitional  epithelium. 

The  bladder  of  tlie  frog  has  already  been  alluded  to  (p.  36). 

The  nerve-ganglia  of  the  bladder  and  ureters  are  best  studied  by  the  lemon-juice  and  gold 
method  (p.  xlv). 


N 


90  PRACTICAL   HISTOLOGY. 


THE  SKIN. 

The  skin  consists  of  {ci)  the  epidermis,  {b)  the  coriiim  (cutis  vera  or  true  skin,  with  the  papillae), 
whilst  deeper  down  is  {c)  subcHtaneo7is  tissue  containing  fat-cells. 

The  epidermis  is  made  up  of  several  layers  ;  the  most  superficial  is  ( i )  the  stratum  corneum, 
or  horny  layer,  composed  of  layers  of  flattened  epithelium  about  to  be  thrown  off  It  varies 
in  thickness,  being  thickest  where  there  is  most  pressure,  e.g.  palms  of  the  hands  and  soles  of 
the  feet.  Below  this  is  (2)  the  stratum  lucidum,  a  narrow,  clear,  homogeneous,  easily-recognised 
layer.  In  it  the  epithelial  cells  are  closely  packed  together,  and  each  one  contains  a  flattened 
nucleus.  Then  follows  (3)  the  stratum  granulosum  (Unna),  or  the  '  stratum  of  granular  cells  ' 
(of  Langerhans).  It  is  best  seen  where  the  skin  is  thick,  and  consists  of  two  or  more  layers 
of  spindle-shaped  '  granular '  and  nucleated  cells,  which  become  deeply  stained  with  the 
carmine  of  picrocarmine,  and  also  with  logwood,  so  that  it  forms  a  marked  feature  in  a  stained 
section.  The  '  granular '  appearance  is  due  to  the  presence  of  granules  of  some  albuminoid 
matter,  probably  closely  related  to  keratin.  The  deepest  layer  (4),  rete  mucosum,  or  rete 
Malpighii,  or  stratum  Malpighii,  consists  of  several  layers  of  epithelium  ;  the  deepest  layers 
are  composed  of  small  columnar  cells,  with  oval  nuclei,  whilst  those  of  the  middle  layers  are 
more  or  less  polyhedral,  with  spherical  nuclei.  In  this  layer  is  deposited  the  pigment  in 
coloured  individuals.  In  it  '  prickle  '  cells  are  easily  seen.  This  layer  rests  on  the  true  skin, 
and  also  dips  down  between  the  papillae  of  the  true  skin. 

The  corium  has,  projecting  from  its  superficial  surface,  a  series  of  conical  or  cylindrical 
papillce,  which  vary  in  size  and  number  in  different  localities,  being  most  numerous  where  the 
sense  of  touch  is  most  acute,  e.g.  palms  of  the  hands  and  soles  of  the  feet.  The  papillae  and  the 
remainder  of  the  true  skin  consist  of  bundles  of  fibrous  tissue,  which  interlace  in  a  very  complex 
manner,  so  as  to  form  a  dense  tissue,  and  in  it  are  to  be  seen  connective-tissue  corpuscles 
and  interfascicular  lymph-spaces,  and  in  it  lie  gland-ducts,  blood-vessels,  nerves,  hairs,  &c. 
It  also  contains  many  elastic  fibres,  which  are  most  numerous  in  the  subcutaneous  tissue  and 
fewest  in  the  papillae.  They  are  arranged  with  reference  to  the  surface  of  the  trabeculae  of 
fibrous  tissue. 

The  subcutaneous  tissue  contains  groups  of  fat-cells,  and  between  them  septa  of  fibrous 
tissue. 

The  skin  also  contains  sweat-glands,  and  in  certain  places  sebaceous  glands  and  hair- 
follicles,  and  numerous  nerves  and  their  terminations,  blood-vessels,  and  lymphatics. 

PREPARATION  {a)  Chromic  acid  and  Spirit  Mixture. — Place  small  pieces  of  the  skin  from 
various  parts,  e.g.  from  the  palm  of  the  hand,  fingers,  or  sole  of  the  foot,  and  scalp,  in  the 
above  fluid  for  ten  days,  and  complete  the  hardening  in  spirit.  Make  vertical  sections  with  a 
freezing  microtome,  and  stain  some  with  logwood,  and  mount  them  in  dammar  ;  others  should 
be  stained  with  picrocarmine,  and  mounted  in  Farrant's  solution,  whilst  others  again  are  to  be 


SKIN.  91 

doubly-stained,  one  set  with  picrocarmine  and  logwood  (dammar),  and  another  with  picrocar- 
mine  and  iodine-green  (dammar),  p.  xlvi. 

{b)  Minute  portions  of  skin  may  be  hardened  in  a  quarter  per  cent.  soUition  o{  osniic  acid 
for  twenty-four  hours,  and  then  preserved  in  spirit. 

(f)  Prepare  pieces  of  the  skin  of  a  human  foetus  in  the  same  way  as  {ci),  but  keep  them 
only  one  week  in  the  hardening  fluid. 

{d)  Place  portions  of  foetal  skin  in  ordinary  alcohol. 

(e)  A  rtificial  digestion,  p.  xxxiv. 

SKIN    OF   THE    PALM. 

Vertical  Section.  (Picrocarmine.)  EXAMINATION  (L). — Observe  the  epidermis,  consisting 
of  many  layers  of  stratified  epithelium.  Beneath  this  the  coriiini,  stained  bright  red,  with 
papillae  projecting  into  the  epidermis,  which  dips  down  between  the  papillae.  In  the  deeper 
part  of  the  skin  note  the  distinct  passage  of  the  connective  tissue  into  that  of  the  subcutane- 
ous tissue,  which  latter  often  contains  masses  of  fat.  Perhaps  the  duct  of  a  sweat-gland  may 
be  seen  running  vertically  in  the  corium,  and  terminating  upwards  in  a  passage  which  winds 
its  way  in  a  corkscrew-like  manner  through  the  epidermis.  In  the  subcutaneous  tissue  will 
be  found  sections  of  the  gland,  where  it  is  arranged  in  the  form  of  a  coil.  Study  these  parts. 
{Indicate  the  general  arrangement  in  one  half  of  VX.  XXI.,  Fig.  i.) 

(H).  Epidermis. — Observe  the  superficial  squames,  stained  yellow,  occurring  in  many  layers 
(stratum  corneum).  They  do  not  appear  to  contain  a  nucleus,  are  flattened,  and  seen  on  edge. 
Beneath  this  note  a  clear  layer,  which  does  not  stain  well,  and  in  which  the  outline  of  the 
cells  is  difficult  to  make  out— the  stratum  lucidum.  Immediately  below  this  there  lies  a  layer 
of  granular  cells  two  or  three  deep,  which  are  deeply  stained  with  carmine,  and  so  stand  out 
brightly  — the  stratum  granulosum  (I^angerhans).  Below  this  observe  the  layers  of  more  or 
less  polyhedral  cells,  with  their  nuclei  stained  red.  Their  edges  often  show  prickles.  This 
layer,  with  those  cells  that  lie  below  it,  constitutes  the  rete  or  stratum  Malpigliii.  The  layer 
of  cells  resting  directly  on  the  corium  is  composed  of  small  columnar  cells  with  oval  nuclei. 
They  seem  to  be  devoid  of  a  membrane.  Compare  the  section  of  the  lip  (p.  60)  for  pigment 
in  these  cells.  {Indicate  these  layers  of  epithelium  in  PL  XXL,  Fig.  2.  Their  relative  position 
is  indicated  by  letters?) 

These  details,  and  especially  the  staining  of  the  nuclei  with  the  carmine,  are  better  seen 
after  the  preparation  has  been  kept  for  a  week.  The  nuclei  gradually  absorb  the  surplus  dye 
from  the  Farrant's  solution. 

Corium  (H). — Observe  the  papillae  and  the  rest  of  the  true  skin,  all  stained  of  a  deep  red 
colour.  Note  the  red-stained  bundles  of  connective  tissue  running  and  interlacing  in  every 
direction,  and  amongst  them  notice  the  large  number  of  elastic  fibres  (stained  yellow)  arranged 
iri  the  form  of  a  network  outside  these  bundles.  Trace  the  continuity  of  the  connective-tissue 
bundles  of  the  corium  with  those  of  the  subcutaneous  tissue.  Here  and  there  in  the  papillae 
capillary  loops  of  blood-vessels,  and  perhaps  nerves,  may  be  seen.%  {Fill  in  papillcc  in  PL  XXL, 
Fig.  2.) 

Sweat-gland  (H). — Observe  the  gland-coil  lying  in  the  subcutaneous  tissue.  Note  the 
gland-tube,  cut  in  all  directions.  Each  tube  consists  of  a  ba.sement-membrane,  lined  by  low 
columnar  or  cubical  cells.  Trace  a  gland-tube  upwards  through  the  corium.  It  consists  of  a 
basement-membrane,  lined  with  low  columnar  epithelial  cells,  which  are  continuous  with,  and 
are  an  extension  of,  the  cells  of  the  stratum  Malpighii.     In  all  cases  the  gland  passes  upwards 

N  2 


92  PRACTICAL   HISTOLOGY. 

between  two  papilla;.  From  the  epidermis  it  pursues  a  spiral  course.  Each  sweat-gland 
consists  of  a  simple  tube,  composed  of  a  coiled  portion — the  gland  proper — situated  in  the 
subcutaneous  tissue,  and  a  duct,  which  passes  more  or  less  vertically  through  the  corium,  to 
open  on  the  surface  of  the  epidermis.  A  brightly  refracting  membrane  seems  to  line  the 
duct  in  its  passage  through  the  epidermis.  In  the  corium,  the  duct  consists  of  a  mcmbrana 
propria,  and  inside  this  are  two  or  three  layers  of  polyhedral  nucleated  cells,  which  are 
separated  from  the  comparatively  narrow  lumen  by  a  homogeneous  bright  membrane.  In  the 
coiled  part,  the  tube  (proximal)  is  lined  with  similar  cells,  but  in  the  other,  or  distal  part,  the 
membrana  propria  is  very  thin,  and  it  is  Hned  with  a  single  layer  of  columnar  nucleated  cells, 
which  are  striated  longitudinally.     {Indicate  a  section  of  sweat-glands  in  PI.  XXL,  Fig.  3.) 

Touch-corpuscles  (Wagner)  may  be  found  in  some  of  the  papillae.  They  are  oval  bodies, 
made  up  of  fibrous  tissue,  and  form  one  of  the  '  end  organs,'  or  structures  in  which  the  nerves 
of  the  skin  terminate. 

In  the  subcutaneous  layers  sections  of  Pacinian  corpuscles  may  be  found  (p.  46),  but  thcj- 
will  be  more  easily  found  in  the  foetal  skin.    {Indicate  a  section  of  one  in  PI.  XXL,  Figs,  i  and  3.) 

DOUBLE-STAINING    OF   THE    SKIN. 

This  is  a  most  excellent  method  for  revealing  the  different  structures. 

1.  Picrocarniine  and  Logivood. — Make  vertical  sections  of  the  skin  of  the  heel  of  a  foetus, 
and  stain  it  first  with  picrocarmine.  Wash  the  section  in  water  slightly  acidulated  with  acetic 
acid,  and  then  wash  it  thoroughly  in  ordinary  water.  Stain  it  slightly  in  logwood  and  mount 
in  dammar. 

The  superficial  layers  of  the  epidermis  are  yellow,  the  deep  layers  and  sweat-glands  are  a 
logwood  tint,  and  the  connective  tissue  is  red. 

2.  Picrocarmine  and  Iodine-green. — Proceed  as  before,  but  use  a  very  dilute  solution  of 
iodine-green  (p.  xliv)  instead  of  the  logwood.     Mount  in  dammar. 

The  superficial  layers  of  the  epidermis  are  yellow,  the  deep  layers,  the  sweat-glands,  nuclei 
of  the  fat-cells  and  connective-tissue  corpuscles  are  green,  whilst  all  the  connective  tissue  is 
red.  These  doubly  and  trebly  stained  sections  are  very  instructive.  In  these  sections  one 
is  almost  sure  to  meet  with  numerous  sections  of  Pacinian  corpuscles  (p.  46).  Note  also 
the  relatively  larger  number  of  sweat-glands  in  the  foetal  skin  (compare  p.  27). 

For  the  development  of  fat-cells  see  p.  27. 


DIGESTION    OF    THE   SKIN. 

This  method  was  introduced  into  histology  several  years  ago  by  the  author.  It  is  invalu- 
able for  ascertaining  the  arrangement  of  the  elastic  fibres  and  muscular  tissue  in  the  skin.  It 
has  recently  been  adopted  by  Kiihne  for  investigating  the  structure  of  nerves.  It  depends  for 
its  value  on  the  fact  that  certain  substances  are  digested  more  rapidly  than  others,  and  so  are 
rapidly  removed. 

Make  an  artificial  gastric  juice  by  mixing  i  c.c.  of  pure  hydrochloric  acid  with  500  c.c. 
water,  and  add  one  gramme  of  pepsine,  or  a  few  drops  of  a  glycerine  extract  of  the  gastric 
mucous  membrane.  It  is  well  to  keep  the  mixture  at  38°  C.  for  two  or  three  hours  before 
using  it.     The  piece  of  skin  to  be  digested  is  stretched  over  a  small  glass  ring  and  firmly  tied 


Plate  X>a.  Skin 


I^-Cntei-rv  Bros  li'-.h 


SKIN.  93 

to  it.  It  is  then  placed  in  some  (200  ex.)  of  the  digesting  fluid,  which  is  kept  at  a  temperature 
of  38°  C.  in  an  ordinary  water  bath  for  a  period  varying  from  three  to  eight  hours, — the  time 
depending  upon  the  age  and  size  of  the  piece  of  slvin.  After  partial  digestion  the  skin  is 
placed  in  water  for  twelve  hours,  when  it  swells  up  and  becomes  extremely  transparent.  It 
may  be  kept  most  advantageously  in  a  ten  per  cent,  solution  of  common  salt,  or  it  may  be 
hardened  in  one  of  the  ordinary  hardening  fluids,  and  afterwards  stained  with  logwood  or 
carmine. 

This  method  is  also  applicable  to  other  tissues  (W.  Stirling). 

HUMAN    SCALP. 

Vertical  Section.  EXAMINATION  (L). — Make  the  sections  parallel  with  the  hair-folliclcs. 
Stain  a  section  with  picrocarmine  or  logwood,  and  mount  it  in  dammar.  Observe  the 
epidermis  and  corium,  and  in  the  latter  the  Iiair-follicles,  each  of  which  gives  origin  to  a 
hair.  Note  its  mouth,  and  its  bulbous  lower  e>itremity,  into  which  projects  a  vascular  papilla. 
The  follicles  are  usually  placed  obliquely,  and  in  the  scalp  they  lie  in  groups  of  three  or 
four,  as  is  easily  seen  when  a  horizontal  section  is  made.  Into  the  upper  part  or  neck  of 
the  follicle  one  or  two  sebaceous  glands  open.  Rising  a  short  distance  above  the  bulbous 
portion  is  the  arrector pili,  composed  of  several  bundles  of  non-striped  muscle-cells.  It  runs 
obliquely  upwards  through  the  corium,  forming  an  acute  angle  with  the  follicle,  and  passes 
over  the  base  of  the  sebaceous  gland,  and  terminates  in  a  plexus  on  the  surface  of  the  corium. 
{Indicate  a  hair-follicle  and  its  muscle  in  PL  XXII.,  Fig.  i.) 

Study  a  Hair-Follicle  (L  and  H). — Proceeding  from  without  inwards,  observe  the  (i)  outer 
and  (2)  inner  siteatk  of  the  hair-sac,  which  are  continuous  with  the  papillary  layer  of  the 
corium.  In  the  outer  layer  the  fibres  are  arranged  longitudinally,  whilst  in  the  inner  one  they 
are  transverse  or  oblique.  Both  layers  are  best  marked  at  the  lower  bulbous  portion  of  the 
follicle,  where  they  are  pushed  into  the  lower  end  of  the  hair,  to  form  a  club-shaped /tf////rt. 
Internal  to  these  tw^o  layers  is  (3)  2.  glassy  hyaline  mevibrane,  which  separates  them  from  the 
epidermic  coverings  of  the  hair.  Inside  this  is  (4)  the  outer  root-sheath,  which  is  thickest  about 
the  middle  of  the  follicle,  and  consists  of  several  layers  of  cells  continuous  with  the  stratum 
Malpighii.  Inside  this  is  (5)  the  inner  root-sheath,  which  in  longitudinal  sections  appears  as 
a  thick  glassy  membrane,  though  it  can  be  proved  to  consist  of  three  layers,  being  from  with- 
out inwards — [a)  Henle's  layer,  then  {b)  Huxley's  layer,  and  {c)  the  cuticle  of  the  root-sheath, 
which  is  a  very  delicate  membrane  composed  of  imbricated  non-nucleated  scales.  The  inner 
root-sheath  terminates  abruptly  at  the  neck  of  the  follicle.  Then  follows  (6)  the  cuticle  of  the 
hair  and  (7)  the  hair  itself. 

Sebaceous  Gland  (L  and  H). — Observe  the  duct  of  one  or  more  of  these  glands  openino- 
into  the  upper  part  of  a  hair-follicle.  The  membrana  propria  (of  the  duct)  is  continuous  with 
the  glassy  membrane  (3)  of  the  hair-sac,  and  the  epithelium  of  the  outer  root-sheath  is  directly 
continuous  with  the  epithelium  of  the  duct.  The  duct  after  branching  opens  into  flask-shaped 
or  saccular  alveoli.  In  these  alveoli  are  found  one  or  more  layers  of  cells  loaded  with  fat- 
globules  (PI.  XXII.,  Fig.  2). 

Arrector  pili  Muscle  (L  and  H). — Observe  its  attachment  to  the  lower  part  of  the  hair- 
follicle  and  trace  it  obliquely  upwards  towards  the  surface  of  the  corium  (p.  91).  It  is  com- 
posed of  non-striped  muscle. 

Horizontal  Sections  of  the  Scalp. — Stain  it  with  logwood  or  picrocarmine,  and  mount  it  in 
dammar.     Observe  the  transverse  sections  of  the  hairs  arranged   in  groups  of  three  or  four. 


94  PRACTICAL   HISTOLOGY. 

Make  out  the  different  layers  of  the  hair-follicle.  Compare  also  the  section  of  a  lip  of  a 
dog  (p.  60)  where  these  various  coverings  are  exquisitely  shown  in  sections  of  the  large 
'  feelers.' 

HAIR. 

Human  Hair.— Pluck  a  hair  from  the  head,  and  place  its  shaft  in  water.  Observe  (H)  its 
cuticle,  consisting  of  imbricated  scales,  then  the  cortex,  which  forms  the  mass  of  the  hair  and  is 
composed  of  horny  matter  made  up  of  scales,  which  in  pigmented  hairs  have  fine  granules  of 
pigment  deposited  between  them.  In  the  centre  note  the  medulla,  or  pith,  which  is  absent  in 
some  hairs  (PI.  XXII.,  Figs.  3  and  4).  A  hair  may  be  preserved  dry,  or  simply  by  adding  a 
drop  of  dammar  solution  and  then  applying  a  cover-glass,  but  after  the  addition  of  dammar 
the  cuticle  is  not  well  seen. 

Hair  of  Rabbit.— Treat  it  similarly,  but  note  the  single  or  double  row  of  oblong  compart- 
ments in  the  medulla  filled  with  air,  and  therefore  appearing  black. 

Transverse  sections  of  hairs  are  obtained  by  shaving,  but  the  hairs  of  the  beard  are  usually 
flattened  ovals,  whilst  those  of  the  head  are  round. 


NAILS. 

Make  vertical  sections  of  a  nail  and  its  nail-bed  after  hardening  in  alcohol  or  chromic  acid 
and  spirit  mixture.  They  are  only  peculiar  modifications  of  the  stratum  Malpighii.  They 
rest  on  very  vascular  papillae  (bed  or  matrix). 

By  steeping  them  in  forty  per  cent,  caustic  potash  solution,  the  nails  can  be  resolved  into 
their  constituent  cells  (p.  xxxiv). 


BLOOD-VESSELS    OF    THE    SKIN. 

PREPARATION. — It  is  best  to  employ  the  skin  of  the  extremities,  for  a  limb  is  easily  in- 
jected from  its  main  artery.  Inject  a  limb  with  a  carmine-gelatine  or  Berlin-blue  mass. 
Harden  parts  of  the  skin  in  Mijller's  fluid  for  two  weeks,  and  then  in  alcohol.  Make  mode- 
rately thick  vertical  sections.  Stain  the  sections  injected  with  a  red  mass  with  logwood,  and 
the  others  with  picrocarmine.  Mount  them  in  dammar.  It  is  well  to  prepare  similar  sections 
of  an  injected  scalp,  to  see  the  arrangement  of  the  blood-vessels  round  a  hair-follicle. 

EXAMINATION  (L  and  H). — Passing  from  the  subcutaneous  tissue  upwards  we  find  the 
following  systems  (Tomsa) : — 

(i)  Observe  the  fat-lobules,  each  supplied  by  a  small  artery  which  forms  a  dense  plexus 
over  and  between  the  individual  fat-cells,  and  from  it  emerge  one  or  two  veins. 

(2)  A  branch  proceeds  upwards  to  the  coil  of  a  sweat-gland,  where  it  forms  a  plexus, 
whilst  the  duct  is  supplied  by  a  branch  from  an  artery  in  the  corium  where  the  capillaries 

anastomose. 

(3)  Each  papilla  of  a  hair-follicle  receives  an  arteriole,  which  splits  up  into  capillaries 
within  it.  A  plexus  of  capillaries  exists  between  the  outer  longitudinal  and  the  inner  circular 
coats  of  the  hair-sac. 

(4)  A  capillary  plexus  exists  round  the  sebaceous  gland  and  arrector  pili. 


Plj^je  xm .  Skin  &  Hair 


""X.,/---     / 


— \^y'    \  ^--'''~\     /""'^  ./''""X..--'" 


MmLei-u  Bros.nU:\. 


SKIN.  95 

(5)  Trace  an  artery  arising  from  a  subcutaneous  trunk,  and  note  that  as  it  ascends  it 
gives  off  branches  to  the  sweat-duct,  hair-sac,  sebaceous  glands,  and  hair-muscle,  and  note  its 
termination  in  capillary  loops  in  the  papillae,  from  which  a  vein  proceeds  which  opens  into 
relatively  large  veins  lying  in  the  superficial  layers  of  the  corium  (Klein).  {Indicate  the 
general  arrangement  of  the  blood-vessels  in  PI.  XXL,  Fig.  i.) 


LYMPHATICS   OF    THE   SKIN. 

They  require  to  be  injected  by  the  puncture  method  (p.  Iv),  using  a  watery  solution 
of  Berlin-blue.  But  the  task  is  by  no  means  an  easy  one.  Injection  of  the  skin  of  the  palm 
of  the  hand  or  sole  of  the  foot  yields  the  best  results. 


NERVES    OF    THE    SKIN. 

The  nerves  of  the  skin  are  very  numerous,  and  the  mode  of  termination  of  some  of  them  in 
Pacinian  corpuscles  in  the  subcutaneous  tissue  (p.  46),  and  in  Wagner's  tactile  corpuscles  in 
some  of  the  papillae  of  the  corium  (p.  92),  has  already  been  alluded  to.  These  touch-cor- 
puscles are  most  numerous  in  the  lips,  skin  of  the  hand,  foot,  and  glans  penis. 

It  requires  some  modification  of  the  gold  method — preferably  that  with  lime-juice  and 
formic  acid  (p.  xlv) — to  obtain  preparations  of  the  various  nerve-plexuses  (sub-epithelial  and 
inter-epithelial)  in  the  skin. 

The  snout  of  the  pig  and  the  skin  of  the  nose  of  the  mole  may  be  employed. 

The  peculiar  nerve-endings  recently  described  by  Ranvier  and  Merkel,  may  be  found  in 
the  snout  of  the  pig,  and  especially  in  the  soft  part  of  a  duck's  bill. 


96  PRACTICAL   HISTOLOGY. 


THE  NERVOUS  SYSTEM. 
THE    SPINAL   CORD. 

PREPARATION  (a).  Miiller's  Fluid  and  Spirit  Mixture. — Remove  the  spinal  cord  and 
medulla  oblongata,  with  the  pia  mater  attached,  from  a  newly-killed  cat,  rabbit,  or  dog. 
Indeed,  all  the  membranes  may  be  left  attached  to  the  cord.  Take  care  to  avoid  squeezing 
or  stretching  the  cord.  It  is  not  necessary  to  make  any  incisions  into  the  cord  at 
first.  Suspend  it  in  a  wide,  cylindrical  vessel — such  vessels  as  are  used  for  water  analysis  do 
very  well — filled  with  a  mixture  composed  of  three  parts  of  Miiller's  fluid,  and  one  of  methylated 
spirit.  The  mixture  ought  to  be  made  at  the  time  ;  but  as  a  considerable  amount  of  heat  is 
evolved  when  the  fluids  are  mixed,  it  ought  to  be  allowed  to  cool.  Keep  the  tissues  in  a  cool — 
in  fact,  a  cold  place.  This  is  of  great  importance.  Renew  the  fluid  at  the  end  of  twenty-four 
hours,  and  then  again  at  the  end  of  a  week.  The  cord  may  now  be  cut,  with  a  sharp  razor,  into 
segments  an  inch  or  so  long.  The  cord  ought  to  be  hardened  in  this  mixture  for  three  weeks. 
It  is  then  transferred  to  a  two  per  cent,  solution  of  ammonium  bichromate  for  two  weeks,  to 
complete  the  hardening.  Preserve  them  either  in  spirit,  or  in  a  solution  of  chloral  hydrate, 
of  twelve  grains  to  an  ounce  of  water  (Hamilton).  A  portion  of  the  spinal  cord  of  a  man  or 
an  ox  ought  to  be  hardened  in  the  same  way. 

{b)  Potassic-bichromate. — Cut  the  fresh  spinal  cord  of  an  ox  into  pieces  an  inch  long  by 
means  of  a  sharp  razor.  Place  them  in  a  large  quantity  of  two  per  cent,  solution  of  potassic 
bichromate.  Do  not  let  one  piece  lie  on  another.  See  that  they  are  separated  by  cotton 
wool,  previously  dipped  in  alcohol,  to  avoid  pressure.  It  requires  from  three  to  five  weeks  to 
complete  the  hardening.  Preserve  the  cord  in  spirit.  A  cord  hardened  in  this  way  shows  the 
nerve-structures  very  well. 

(c)  Chromic  Acid  and  Spirit  Mixture. — The  cord  must  be  cut  into  pieces  half  an  inch  long 
and  placed  in  this  fluid,  which  will  harden  them  in  from  two  to  three  weeks.  It  is  not  so  well 
suited  for  showing  the  nerve-elements  as  for  demonstrating  the  neurogleia. 

id)  Spirit  and  Iodine,  and  then  Ammonium-bichromate. — Suspend  the  cord  in  a  long,  cylindrical 
vessel,  in  methylated  spirit  tinged  yellow  with  tincture  of  iodine.  Leave  it  in  this  mixture 
— usually  three  or  four  days — until  the  yellow  colour  has  nearly  disappeared.  Cut  it  in  pieces 
an  inch  long,  and  complete  the  hardening  by  keeping  them  in  a  two  per  cent,  solution  of  am- 
monium-bichromate for  four  or  five  weeks.     Preserve  them  in  spirit  until  required  for  section. 

METHOD  OF    CUTTING   SECTIONS  OF   THE    SPINAL   CORD. 

Employ  some  form  of  freezing  microtome.  Steep  a  piece  of  the  cord,  hardened  by  any 
of  the  above  methods,  but  preferably  by  («),  in  water  for  twenty-four  hours,  to  remove 
all  traces  of  the  hardening  agent,  or  the   spirit.     Soak  it  in  syrup  (two  ounces  of  sugar  to 


SPINAL    CORD.  97 

one  ounce  of  water)  for  t\vcnt)--four  or  thirty-six  hours,  and  then  transfer  it  to  a  solution  of 
gum  for  twelve  hours,  and  then  cut  it  in  the  ordinary  way  with  a  freezing  microtome. 

Stain  the  sections  with  logwood,  carmine,  or  aniline  blue-black,  and  mount  them  in 
dammar. 

When  investigating  the  structure  of  the  nervous  system,  it  is  sometimes  of  importance 
to  have  a  section  only  partial/)'  cleared  up.  This  may  be  done  by  examining  a  section  in 
Farrant's  solution,  though,  for  general  purposes,  mounting  in  dammar  is  to  be  preferred. 

Make  transverse  sections  through  the  hardened  cervical  portion  of  the  cord  of  a  cat,  rabbit, 
or  dog.  Stain  a  section  with  carmine,  or  logwood,  or  preferably  with  aniline  blue-black  one 
per  cent,  solution,  which  stains  the  nerve-cells  beautifully  (p.  xliv).  Mount  the  sections  in 
dammar. 

Transverse  Section  of  the  Spinal  Cord.  EXAMINATION  (L). —  Observe  the  pia  mater 
investing  the  cord.  It  consists  of  an  outer  part  and  an  inner  part,  which  latter  sends  numerous 
processes  or  septa  into  the  cord.  Trace  the  passage  of  the  outer  layer  into  the  anterior 
median  fissure,  and  a  process  of  the  inner  layer  into  the  posterior  median  fissure. 

Observe  that  the  cord  is  essentially  a  bilateral,  symmetrical  organ,  its  two  halves  being 
connected  by  a  narrow  bridge  of  tissue  containing  a  small  opening — the  central  canal.  Note 
the  broad,  shallow,  and  well-defined  anterior  median  fissure,  and  the  narrow,  deeper,  and  less 
perfectly  de?meA  posterior  median  fissure,  containing  a  process  of  the  pia  mater,  often  carrying 
a  blood-vessel. 

If  the  section  has  been  made  at  the  level  of  the  origin  of  the  nerve-roots,  trace  the  fibres  of 
the  anterior  roots  of  a  spinal  nerve,  leaving  the  cord  by  several  bundles,  whilst  those  of  the 
posterior  root  enter  it  as  one  bundle. 

White  matter. — Observe  the  white  matter,  composed  of  the  cut  ends  of  nerve-fibres  of 
the  cord  placed  externally,  and  how  each  half  is  divided  into  an  anterior,  lateral,  and  posterior 
column  by  the  above-described  arrangement  of  the  nerve-roots.  The  posterior  column  is  ao-ain 
divided  into  two  (in  the  cervical  region),  by  a  narrow  band  of  connective  tissue  ;  the  narrow 
inner  part,  which  lies  next  the  posterior  median  fissure,  is  known  as  the  fasciculus  cuneatus. 

Grey  Matter. — Observe  the  grey  matter  more  deeply  stained,  and  lying  in  the  form  of  a 
crescent  in  each  half  of  the  cord.  Its  extremities  constitute  the  anterior  and  posterior  horns 
or  cornua,  though  there  is  no  well-marked  limit  between  them.  Note  in  the  anterior  horn 
the  numerous  multipolar  nerve-cells  deeply  stained,  and  observe  the  oval  cap  of  deeply 
stained  matter — the  substantia  gclatinosa — which  covers  the  posterior  cornua.  It  is  probably 
non-nervous  in  its  nature,  and  consists  of  a  peculiar  accumulation  of  '  neurogleia.'  Observe 
the  bridge  of  tissue  connecting  the  two  halves  of  the  cord,  and  in  it  the  central  canal  lined 
with  columnar  ciliated  epithelium, — in  front  and  behind  the  canal,  the  anterior  and  posterior 
grey  commissures,  also  the  nerve-fibres  crossing  in  front  of  the  anterior  grey  commissure, 
forming  the  anterior  white  commissure.  {Indicate  the  general  arrangement  in  PI.  XXIII. 
Fig.  I.)  ■ 

White  Matter.  (H). — Observe  the  pia  mater  sending  in  septa  ;  trace  these  septa,  becomino- 
continuous  with  a  fine  fibrous  network — the  neurogleia — lying  between  the  nerve-fibres  of  the 
white  substance.  Lying  in  the  meshes  of  this  network  observe  the  cut  ends  of  the  nerve-fibres. 
They  are  of  different  sizes  :  some  are  large,  others  of  medium  size,  and  some  are  very  small. 
Each  axis-cylinder  is  deeply  stained,  and  is  surrounded  by  a  clear,  non-stained  area,  which 
represents  the  white  substance  of  Schwann,  or  myeline.  In  sections  which  have  been 
hardened  for  a  long  time  in  chromic  acid,  a  well-marked  concentric  striation  of  the  medulla 

O 


98  PRACTICAL  HISTOLOGY. 

may  be  observed  (see  Nerve,  p.  42).  {Indicate  the  nervc-fibrcs  in  one  half  and  the  ncurogleia  in 
the  other  half  of  Vl  XXIII.,  Fig.  2.) 

Ncurogleia  (H). — This  is  the  deHcate  framework  which  supports  the  nerve-elements — fibres 
and  cells — within  the  cord  and  brain.  It  fills  all  the  interstices  between  the  nervous  elements 
of  the  cord.  It  consists  of  a  semi-fluid  substance — the  neurogleia  matrix — in  which  are 
imbedded  numerous  very  fine  neurogleia  fibrils,  which  unite  to  form  a  network.  In  it  are 
also  found  many  nucleated,  branched,  connective-tissue  corpuscles.  The  fibres  seem  to  be 
allied  in  their  nature  to  clastic  fibres. 

Grey  Matter  (H). — Observe  the  large,  nucleated,  multipolar  nerve-cells  in  the  anterior  horn  ; 
four  or  five  branching  processes  may  be  traced  from  them  for  a  considerable  distance.  Each 
cell  contains  a  distinct  spherical  nucleus,  enclosed  in  a  distinct  membrane  and  a  nucleolus. 
Round  each  cell  is  a  space— the  lymph-space.  These  cells  are  aggregated  into  three  groups — 
an  anterior,  lateral,  and  median.  Study  the  posterior  horn,  and  notice  that  the  cells  there  are 
much  smaller  in  size  and  fewer  in  number.  Notice  the  fibrillar  arrangement  of  the  rest  of  the 
grey  matter,  and  study  the  substantia  gelatinosa  of  Rolando  (p.  97),  which  is  composed  of 
a  mass  of  neurogleia.     {Indicate  the  anterior  horn,  with  its  nerve-cells,  in  PI.  XXIII.,  Fig.  3.) 

Study  similar  sections  made  from  the  dorsal  and  lumbar  regions  of  the  cord,  and  compare 
them  with  the  above.  Observe  that  the  sections  of  the  dorsal  region  are  smaller  than  the 
others.  Compare  the  absolute  amount  of  grey  matter,  and  observe  that  it  is  greatest  in  the 
lumbar  and  cervical  regions.  The  amount  of  white  matter  is  least  in  the  lumbar  and  dorsal 
regions,  and  greatest  in  the  cervical.  In  the  upper  dorsal  region  notice  a  group  of  cells  in  the 
grey  matter,  midway  between  the  anterior  and  posterior  cornua— the  tractus  inter-niedio- 
lateralis. 

Make  similar  sections  of  the  spinal  cord  of  an  ox  or  man,  or,  best  of  all,  that  of  a  horse. 

It  is  necessary  to  trace  the  course  of  the  fibres  as  seen  in  a  section.  Observe  (L)  the 
atiterior  root  of  a  spinal  nerve.  Trace  some  of  its  fibres.  They  enter  the  anterior  cornu 
of  its  own  side,  and  some  of  the  fibres  pass  through  the  anterior  white  commissure  to  the 
anterior  column  of  the  opposite  side.  Others  turn  into  the  lateral  column  on  the  same  side. 
Trace  the  fibres  of  the  posterior  root  entering  as  a  single  bundle  through  the  substantia 
gelatinosa. 

To  study  further  the  course  of  the  nerve-fibres  it  is  necessary  to  make  antero-posterior 
sections  through  the  grey  and  white  matter.  This  is  best  done  in  the  cervical  region  of  the 
cord  of  an  ox.     Stain,  and  mount  in  the  same  way. 

Longitudinal  Section  of  the  Spinal  Cord.  EXAMINATION  (L).— Observe  the  grey  matter  in 
the  centre  with  a  band  of  white  matter— the  anterior  and  posterior  columns  cut  longitudinally- - 
on  each  side  of  it.  Trace  the  vertically-arranged  nerve-fibres  in  these,  and  note  the  some- 
what oblique  entrance  of  the  anterior  roots,  some  of  the  fibres  ascending,  and  others  descending 
in  the  grey  matter.  In  the  anterior  cornu  note  the  groups  of  multipolar  nerve-cells  and  also 
the  gelatinous  substance  in  the  posterior  column  and  the  entrance  of  a  posterior  root. 

TO    ISOLATE   A   MULTIPOLAR    NERVE-CELL   OF   THE    SPINAL   CORD. 

PREPARATION,  {a)  Dilute  Potassic  Bichromate. — Macerate  a  small  piece — an  eighth  of 
an  inch  long— of  the  perfectly  fresh  spinal  cord  of  an  ox  or  sheep  in  a  large  quantity  of  a 
one-eighth  per  cent,  solution  of  potassic  bichromate  for  two  or  three  days.  After  this  time, 
cut  out  with  scissors  a  small  piece  of  the  anterior  horn,  and  tease  it  on  a  slide  in  water.  Stain 
the  piece  with  carmine,  and  then  continue  to  tease  it  with  needles  until   isolated  cells  are 


Plvje  X>[E.  Spih^  Cord 


0 


SPINAL    CORD.  99 

obtained.  The  process  of  teasing  is  best  accomplished  with  the  aid  of  a  dissecting  microscope, 
but  the  preparation  ought  to  be  examined  from  time  to  time  with  a  low  power.  A  hair  or 
thin  piece  of  paper  ought  to  be  placed  under  the  cover-glass,  to  prevent  the  cells  from  being 
displaced. 

(b)  Injection  of  Osmic  Acid. — By  means  of  a  subcutaneous  syringe,  provided  with  a  very  fine 
gold  nozzle,  inject  forcibly  a  small  quantity  of  a  half  per  cent,  solution  of  osmic  acid  into  the 
anterior  horn  of  a  perfectly  fresh  spinal  cord.  This  serves  to  separate  the  parts,  and  at  the 
same  time  to  fix  them.  Macerate  the  part  for  two  days  in  dilute  alcohol,  and  isolate  the  cells 
by  carefully  teasing  out  a  piece,  as  described  under  {a). 

(c)  Dilute  Osmic  Acid. — Macerate  pieces,  the  size  of  half  a  pea,  of  the  grey  matter  of  the 
anterior  horn  of  a  perfectly  fresh  cord  in  a  considerable  quantity  of  a  tenth  per  cent,  solution 
of  osmic  acid  for  ten  days.  Wash  away  the  greyish  deposit,  and  place  the  pieces  in  a  mixture 
consisting  of  equal  parts  of  glycerine  and  water,  for  ten  days  or  a  fortnight.  Tease  a  small 
piece  in  glycerine.     The  cells  are  easily  stained  with  a  very  dilute  solution  of  magenta. 

EXAMINATION  (L  and  H). — Observe  the  large  multipolar  cells,  with  many  branched 
processes.  One  process — the  axis-cylinder  process,  which  is  directly  continuous  with  the 
axis-cylinder  of  a  nerve-fibre — is  always  unbranched.  Observe  the  large  nucleus,  and  note  the 
longitudinal  striation  of  the  cell-substance  and  that  of  the  axis-cylinder  process. 

BLOOD-VESSELS    OF    THE    SPINAL   CORD. 

An  entire  animal  must  be  injected,  and  sections  made  of  the  injected  cord.  The  grey 
matter  is  much  more  vascular  than  the  white.  The  sections  are  mounted  in  dammar,  and 
need  not  be  stained. 

The  connective  tissue  of  the  subarachnoid  space  has  already  been  referred  to  (p.  23). 


THE    MEDULLA   OBLONGATA. 

PREPARATION. — The  same  methods  as  are  employed  for  the  spinal  cord  are  to  be  used. 
Make  transverse  sections  of  the  human  medulla,  some  through  the  decussation  of  the  anterior 
pyramids,  and  others  through  the  floor  of  the  medulla  and  the  olivary  bodies.  Stain  them 
with  aniline  blue-black,  and  mount  in  dammar. 

EXAMINATION  (L).  Transverse  Section  through  the  decussation  of  the  Pyramids.—  Observe 
the  crossing  of  the  fibres  from  one  side  to  the  other  ;  notice  the  enlarged  posterior  cornua 
(tubercles  of  Rolando),  the  central  canal  nearer  the  surface  posteriorly. 

Through  the  Olivary  Body. — Observe  the  median  raphe  ;  the  central  canal  has  now  ex- 
panded into  the  floor  of  the  fourth  ventricle.  Study  the  columns  of  the  medulla  from  the 
centre  in  front  Outwards  as  anterior  pyramids,  olivary  body,  restiform  body,  and  posterior 
pyramid  on  each  side.  Notice  the  folded  sheet  of  grey  matter — corpus  dentatum — in  the 
olivary  body,  and  in  it  notice  the  multipolar  nerve-cells.  Observe  the  masses  of  grey  matter 
in  the  floor  of  the  fourth  ventricle. 


loo  PRACTICAL   HISTOLOGY. 


THE    CEREBELLUM. 

PREPARATION. — The  same  methods  are  applicable  as  those  detailed  for  the  spinal  cord. 
Harden  pieces — three-quarters  of  an  inch  square — of  a  human  cerebellum  in  the  Miiller's 
fluid  and  spirit  mixture  (p.  xxxii),  and  proceed  just  as  for  the  spinal  cord.  If  a  human 
cerebellum  cannot  be  obtained,  use  that  of  a  cat,  dog,  or  rabbit.  Make  sections  across  the 
folia  or  leaflets.  Stain  some  with  logwood,  but  preferably  with  aniline  blue-black,  and  mount 
them  in  dammar. 

Vertical  Section  of  the  Cerebellum.  EXAMINATION  (L).— Observe  the  folia  or  leaflets,  with 
secondary  leaflets  on  them.  Notice  the  dark-stained  grey  matter  outside,  and  the  lighter 
white  matter  inside.  The  grey  matter  is  covered  with  pia  mater,  which  sends  processes 
carrying  blood-vessels  into  the  substance  of  the  organ.  It  consists  of  (i)  a  broad,  outer  grey 
or  molecular  layer,  containing  at  its  inner  part  a  single  row  of  pear  or  flask-shaped  cells';  (2) 
the  cells  of  Purkinje,  which  give  off  a  number  of  branching  processes,  running  outward  ; 
inside  this  is  (3)  the  inner  granular  or  nuclear  layer,  about  one-third  the  breadth  of  (1). 
Observe  the  large  number  of  small  deeply  stained  granules,  arranged  in  many  rows  ;  and  then 
follows  the  white  matter,  consisting  of  nerve-fibres  arranged  more  or  less  parallel  to  each  other. 
{Indicate  the  layers  in  PL  XXIV.,  Fig.  i.) 

(H).  (i)  The  outer  or  cortical  layer  contains,  as  a  ground-work,  a  network  of  delicate 
nerve-fibrils,  and  mixed  with  these  are  the  dichotomously  divided  branches  of  Purkinje's  cells. 
It  contains  a  few  multipolar  ganglion-cells  (Sankey),  which  seem  to  be  connected  with  these 
processes. 

(2)  Purkinje  s  cells  occur  in  a  single  layer.  They  are  of  a  pear  or  flask-like  shape.  Each 
cell  has  a  single,  unbranched,  or  axial  cylinder  process,  which  passes  centrally  ;  whilst  from 
its  outer  surface  it  gives  off  a  branched  process,  which  splits  up  rapidly  into  branches  or  pro- 
cesses which  divide  again  and  again.     They  can  be  traced  nearly  to  the  surface  of  the  organ. 

(3).  The  nuclear  layer  is  a  broad  layer,  and  contains  a  great  number  of  deeply-stained 
spherical  nuclei,  whose  exact  nature  is  unknown.  Between  them  is  a  network  of  minute 
fibrils,  which   are  probably   nervous    in  their   nature.      {Indicate  these  layers  in  PL  XXIV., 

Fig.  2.) 

In  the  white  matter  note  the  small  nerve-tubes,  and  between  them  the  neurogleia,  con- 
taining rows  of  small  nucleated  cells. 

Trace  a  blood-vessel  in  the  grey  matter,  and  observe  that  each  one  is  surrounded  by  a 
narrow  space — the  perivascular  lymph-space  (His). 

DOUBLE-STAINING   OF   THE    CEREBELLUM. 

(i)  Eosin  and  Logwood. — Stain  a  section  in  a  very  dilute  watery  solution  of  eosin  (p.  xlv) 
till  it  has  a  faint  red  colour.  This  it  does  in  a  few  seconds.  Great  care  must  be  taken  not 
to  overstain  the  preparation.  This  forms  an  excellent  ground-colour.  Wash  the  section  in 
water,  and  then  stain  it  with  logwood,  and  mount  it  in  dammar.  The  cortical  layer  is  reddish, 
and  so  are  the  cells  of  Purkinje,  while  the  nuclear  layer,  and  all  other  nuclei,  have  a  logwood 
tint. 

(2)  Picrocarmine  and  Iodine-green.— Stain  a  section  with  picrocarmine.     Wash  it  in  water 


CEREBRUM. 


lOI 


acidulated  with  dilute  acetic  acid,  and,  after  washing  in  pure  water,  stain  it  with  iodine-^reen, 
and  mount  it  in  dammar.  The  outer  layer  and  Purkinje's  cells  of  the  grey  matter  are  stained 
red,  and  the  inner  layer  is  stained  green. 


THE    CEREBRUM. 

PREPARATION. — Prepare  it  in  the  same  way  as  the  cerebellum  (p.  loo),  using  the  same 
precautions  both  for  hardening  the  tissue  and  for  cutting  it  as  indicated  for  the  spinal  cord 
(p.  96).  Select  a  frontal  convolution  of  a  human  brain  for  examination.  Stain  the  sections 
with  aniline  blue-black,  and  mount  them  in  dammar. 

Vertical  Section  of  a  Human  Cerebral  Convolution.  EXAMINATION  (L). — Observe  the  shape 
of  a  convolution,  and  contrast  it  with  that  of  the  cerebellum.  Notice  the  pia  mater  surround- 
ing the  convolution,  and  sending  fine  processes  carrying  blood-vessels  into  the  cortex.  Notice 
the  deeply  stained  grey  matter  or  cortex,  and  the  white  matter  within.  Examine  the  layers 
of  the  grey  matter  from  without  inwards.  It  is  possible  to  distinguish  five  layers,  varying  in 
thickness  and  structural  characters  (Meynert).  These  layers  are  not  accurately  mapped  off  from 
each  other  ;  still  they  can  be  seen.  It  is  to  be  remembered,  however,  that  there  are  variations 
in  different  parts  of  the  cortex.     In  some  places  only  four  layers  can  be  distinguished. 

1.  The  outer  layer  contains  few  cells  of  any  kind  imbedded  in  a  clear  matrix.  It  occupies 
about  one-tenth  of  the  entire  thickness  of  the  grey  matter. 

2.  This  is  about  equal  in  thickness  to  the  preceding,  though  it  is  easily  distinguished  by  its 
containing  a  large  number  of  densely  packed  small  pyramidal  cells.  Owing  to  the  large 
number  of  cells  in  this  lajer  it  always  appears  deeply  stained. 

3.  This  layer  is  much  wider,  and,  owing  to  the  cells  occurring  in  it  being  placed  well  apart 
from  each  other,  it  appears  somewhat  lighter  than  the  second  layer.  It  consists  of  large  and 
small  pyramidal  cells,  so  placed  that  their  apices  are  directed  towards  the  surface  of  the  con- 
volution, and  their  bases  towards  the  white  matter. 

4.  This  layer  contains  small  irregular  corpuscles  with  few  processes. 

5.  This  layer  is  broader  than  the  one  above  it,  and  contains  similar  corpuscles  mixed  with 
a  few  fusiform  cells.  At  its  lower  part  it  gradually  shades  into  the  wliite  matter,  which  con- 
sists of  very  fine  medullated  fibres,  supported  by  neurogleia.  {Indicate  t lie  general  arrangement 
of  the  cells  in  PI.  XXIV.,  Fig.  3.) 

(H).  Study  in  detail  the  various  layers,  and  especially  note  the  cells  of  the  third  layer. 
The  cells  are  distinctly  pyramidal.  Trace  the  long,  branching  process  directed  towards  the 
surface,  and  the  branching  processes  from  each  angle  of  the  base.  Each  cell  has  a  central  axial 
cylinder-process.  Each  ganglion-cell  lies  in  a  lymph-space.  It  is  to  be  observed,  however, 
that  these  cells  only  appear  pyramidal  when  the  plane  of  the  section  passes  parallel  to  their 
long  axis.  If  cut  transversely  they  appear  triangular.  {Indicate  a  nerve-cell  in  PI.  XXIV., 
Fig.  4-) 

Select  a  blood-vessel  of  the  grey  matter,  and  observe  its  perivascular  lymph-space. 

The  processes  of  the  cells  are  best  seen  in  preparations  which  are  ox\\y  partially  cleared  up 
under  the  influence  of  clove  oil.  This  is  a  most  important  method  of  investigation.  I  have 
often  seen  in  this  way  delicate  fibrils,  not  unlike  elastic  fibres,  and  which  are  not  distinct  when 
the  section  is  completely  cleared  up. 


I02  PRACTICAL   HISTOLOGY. 


BLOOD-VESSELS    OF   THE    BRAIN. 

The  entire  animal  must  be  injected.     Make  vertical  sections  and  mount  them  in  dammar. 
Observe  the  greater  vascularity  of  the  grey  matter. 


NERVE-GANGLIA.    GASSERIAN    OR   SPINAL   GANGLIA. 

PREPARATION. — With  a  saw  divide  longitudinally  the  head  of  a  sheep  just  killed.  With 
a  scalpel  remove  the  Gasserian  ganglia  from  the  apex  of  the  petrous  part  of  the  temporal  bone, 
and  place  them  in  the  chromic  acid  and  spirit  fluid  for  eight  days.  Make  longitudinal  and 
transverse  sections.  Stain  them  with  logwood  and  mount  them  in  dammar,  or  in  Farrant's 
solution.  The  ganglion  on  the  posterior  root  of  a  spinal  nerve  of  a  cat  or  dog  may  be 
hardened  by  the  same  method. 

Longitudinal  Section  of  a  Spinal  Ganglion.  EXAMINATION  (L).— Observe  the  sheath  of  the 
ganglion^within  it  are  numerous  large  ganglionic  cells — and  note  their  arrangement.  Ob- 
serve strands  of  nerve-fibres  sweeping  through  the  ganglion,  and  processes  of  connective  tissue 
passing  in  from  the  capsule  into  the  ganglion,  and  supporting  the  cells  and  nerve-fibres.  {In- 
dicate the  nerve-cells  and  fibres  in  PI.  XXIV.,  Fig.  5.) 

(H).  Observe  the  nerve-cells,  with  their  well-marked  sheath  or  capsule  lined  with  squames, 
the  nuclei  of  which  are  well-marked  ;  the  granular  mass  of  the  cell,  containing  a  round  ex- 
centric  nucleus  with  an  envelope,  and  a  distinct  nucleolus.  It  is  difficult  to  recognise  the 
processes  of  the  cell  itself     Note  the  medullated  nerve-fibres. 

In  the  examination  of  the  transverse  sections  note  the  cells  as  before,  but  between  them 
bundles  of  medullated  nerve-fibres  cut  transversely. 

It  is  also  desirable  to  examine  the  cells  from  a  fresh  Gasserian  ganglion.  For  this  purpose 
tease  a  small  piece  of  the  fresh  Gasserian  ganglion  of  a  sheep  in  a  very  small  quantity  of  salt 
solution,  and  stain  it  with  magenta  solution.  Details  similar  to  those  described  above  are  seen 
on  examination,  though  the  processes  of  the  cells  are  generally  broken  off,  and  the  cells  them- 
selves are  frequently  dislocated  from  their  capsules.  {Indicate  an  isolated  cell  within  its  capsule 
in  PI.  XXIV.,  Fig.  7,  and  without  its  capside  in  Fig.  6.) 

GANGLIA   OF   THE    SYMPATHETIC    NERVOUS    SYSTEM. 

The  non-medullated  nerve-fibres  have  already  been  examined  (p.  45).  The  nerve-cells 
of  the  sympathetic  system  have  still  to  be  considered. 

PREPARATION.— Remove  a  sympathetic  ganglion  from  the  thorax  of  any  animal,  or  pre- 
ferably use  the  human  superior  or  inferior  cervical  sympathetic  ganglion.  Harden  it  in  Miiller's 
fluid,  or  the  chromic  acid  and  spirit  fluid,  for  four  or  five  days,  and  make  transverse  sections. 
Stain  sections  with  logwood,  and  others  with  picrocarmine,  and  mount  them  in  Farrant's 
solution. 

EXAMINATION  (L). — Observe  the  capsule  of  the  ganglion  composed  of  fibrous  tissue,  and 
notice  the  cells  and  their  arrangement.     Place  a  cell  in  the  field  of  the  microscope. 

(H).  Observe  the  cell-capsule,  with  its  included  nucleated  cell,  which  gives  off  processes 


Plate XXiy.  Brain  &,Nerve  Ganglia 


Miiiten-i.  Eros    hth 


NERVE-GANGLIA.  103 

that  pierce    the  capsule.     The  cells  very  frequently   contain  a   yellowish-coloured   pigment. 
Between  the  cells  will  be  found  transverse  sections  of  non-medullated  nerve-fibres. 

SYMPATHETIC    NERVE-CELLS    OF    THE    FROG. 

These  have  already  been  alluded  to  in  connection  with  the  heart.  They  occur  in  large 
numbers  along  the  course  of  the  abdominal  aorta  in  the  frog,  and  are  easily  demonstrated  by 
the  lime-juice  and  gold  method  (p.  xlv).  They  may  also  be  obtained  from  the  sympathetic 
ganglia.  It  is  easy  to  demonstrate  the  straight  process,  but  by  no  means  so  easy  to  see  the 
spiral  one. 

Isolation  of  Sympathetic  Nerve-cells. — Adopt  the  same  methods  as  are  recommended  for 
the  nerve-cells  of  the  spinal  cord. 


I04  PRACTICAL   HISTOLOGY. 


THE    EYE. 
THE    CORNEA. 

PREPARATION. — Cut  out  the  fresh  cornea  from  the  eye  of  a  cat  or  rabbit.  In  excising 
the  eyeball,  be  careful  not  to  squeeze  or  injure  it.  For  the  removal  of  the  cornea,  push  a 
narrow  knife  into  it  at  its  junction  with  the  sclerotic,  and  then  cut  it  out  with  scissors. 
Harden  it  in  a  two  per  cent,  solution  of  potassic  bichromate  for  ten  days,  or  in  the  chromic 
acid  and  spirit  mixture  for  a  week.  Make  vertical  sections,  stain  some  with  logwood  and  others 
with  picrocarmine,  and  mount  in  Farrant's  solution,  and  also  irusdammar. 

EXAMINATION  (L). — Observe  the  epithelium  occurring  in  many  layers — conjunctiva  cornecu 
or  anterior  epithehmn — covering  the  anterior  surface  of  the  cornea  proper.  In  the  human 
cornea,  and  in  the  cornea  of  some  animals,  the  epithelium  rests  on  a  narrow,  elastic,  trans- 
parent membrane — the  anterior  elastic  lamina  or  Bowniati's  membrane — but  it  is  either  absent 
or  but  feebly  developed  in  the  cornea  of  the  cat  and  dog.  It  seems  to  differ  from  the  next 
layer  chiefly  in  the  absence  of  corneal  corpuscles.  The  substance  forming  the  cornea  proper — 
stcbstantia  propria — is  stained  red,  and  on  its  posterior  surface  there  is  a  section  of  a  clear, 
sharply-defined  membrane  stained  yellow — the  membrane  of  Descemet,  or  posterior  elastic 
lamina and  covering  the  posterior  surface  of  this  membrane  there  is  a  single  layer  of  flat- 
tened nucleated  epithelial  cells  seen  in  profile.  Examine  each  of  these  parts.  {Indicate  the 
general  arrangement  in  one  half  of  Y\.  XXV.,  Fig.  i.) 

(H).  The  anterior  epithelium.  Begin  at  the  surface,  and  note  the  flattened  cells  seen  in 
section.  The  deeper  layers  consist  of  two  or  three  layers  of  polyhedral  cells,  each  with  a 
spherical  nucleus.  They  are  so  arranged  as  to  fit  into  each  other,  and  the  deepest  layer  resting 
on  the  cornea  is  compo.sed  of  a  row  of  columnar  cells  placed  perpendicularly.  Observe  how 
processes  of  the  cells  in  the  layer  above  this  fit  into  depressions  between  the  apices  of  the 
columnar  cells  (Cleland  and  Stirling).  The  columnar  cells  are  not  all  of  the  same  height,  and 
their  lower  end  possesses  a  flat  expansion  or  foot-plate  (Lott  and  Stirling),  which  rests  on 
the  cornea.  Prickle-cells  are  seen  in  the  middle  layers,  especially  where  the  epithelium  is 
in  very  many  layers,  as  in  the  cornea  of  the  ox.     (See  p.  9.) 

The  anterior  elastic  lamina  is  a  homogeneous,  transparent  membrane  devoid  of  cornea- 
corpuscles.  It  is  perforated  here  and  there  by  a  few  oblique  channels,  which  transmit  fine 
nerve-fibrils— the  rami  perforantes. 

The  substantia  propria  is  composed  of  a  series  of  layers  of  fibrillar  connective  tissue, 
arranged  in  the  form  of  lamellae,  placed  one  outside  the  other,  and  parallel  to  the  surface  of 
the  cornea.  These  lamella:,  and  also  the  fibres  which  compose  them,  are  held  together  by  an 
albuminous  cement-substance.  Bundles  of  fibrous  tissue  pass  from  one  lamella  to  another. 
Near  the  anterior  surface  a  few  fibres  perforate  the  lamella  obliquely,  and  constitute  the 


CORNEA.  J  05 

fibm  arciiata.  Note  the  edges  of  these  lamellas  seen  in  section.  In  the  ground -substance 
which  separates  two  adjoining  lamcUfe,  there  exists  a  series  of  lacuna;  or  cell-spaces  of 
irregular  branched  form,  which  communicate  freely  by  means  of  fine  channels  with  lacunae  in 
the  same  plane,  and  also  with  the  lacunae  lying  above  and  below  them.  This  is  the  lymph- 
canalicular  system  of  V.  Recklinghausen.  Each  lacuna  contains  a  nucleated  cell-plate  or 
corneal  corpuscle^  which  does  not  completely  fill  the  space,  so  that  lymph  and  colourless  blood- 
corpuscles  can  move  between  it  and  the  wall  of  the  space.  These  corpuscles  are  seen  as  flat, 
narrow,  nucleated,  stained  spindles  lying  between  the  lamellae  in  oval  spaces,  which  they  do 
not  fill  completely.  Each  corpuscle  sends  processes  into  the  canaliculi,  so  that  they  anasto- 
mose to  form  a  network.  They  cannot  be  seen  in  this  preparation  ;  it  requires  the  use  of 
gold  chloride  to  show  their  processes  satisfactorily  (p.  105). 

The  posterior  elastic  lamina  is  a  structureless  elastic  membrane  with  sharpiiy  defined 
margins,  which  stains  of  a  deep  yellow  tint  with  picrocarmine.  It  may  become  detached, 
when  it  curls  up.  It  consists  of  fibrils.  Study  the  single  layer  of  polyhedral,  nucleated, 
granular,  endotlielial  cells  on  its  posterior  surface  {jneinbrana  Dcscemeti). 


CORNEA-CORPUSCLES,  AND  NERVES  OF  THE  CORNEA. 

PREPARATION. — These  are  both  demonstrated  by  the  same  method,  viz.  the  gold  method. 
This  may  be  accomplished  in  several  ways. 

{a)  Reduction  by  Dilute  Acetic  Acid. — Remove  the  cornea  from  an  animal  just  killed  {e.g. 
cat,  rabbit,  or  guinea-pig),  using  the  same  precautions  to  avoid  stretching  it  as  indicated  at 
p.  104.  Place  it  in  a  few  c.c.  of  a  half  per  cent,  solution  of  gold  chloride,  and  keep  it  in  the 
dark.  Let  it  remain  in  the  gold  for  half  an  hour  to  an  hour  in  the  case  of  the  cornea  of  a 
guinea-pig,  and  an  hour  and  a  half  to  two  hours  in  that  of  a  rabbit.  Remove  it,  and  wash  it 
in  distilled  water,  and  place  it  in  water  slightly  acidulated  with  acetic  acid,  and  expose  it  to 
the  light  until  it  becomes  of  a  decidedly  purplish  or  slate-grey  colour  throughout.  Make 
vertical  and  horizontal  sections,  and  mount  them  in  glycerine. 

{b)  Reduction  by  strong  Tartaric  Acid. — Proceed  as  before  up  to  the  stage  where  the  cornea 
is  washed  in  distilled  water.  Expose  it  for  a  day,  or  two  days,  to  the  light  in  distilled  water 
without  the  addition  of  any  acid,  until  it  assumes  a  violet  colour.  Place  it  in  a  mi.xture  of 
glycerine,  one  part,  and  water,  two  parts,  for  two  or  three  days,  and  keep  it  in  the  dark. 
Wash,  and  place  it  in  a  small  beaker  half  filled  with  a  nearly  saturated  watery  solution  of 
tartaric  acid.  Heat  this  in  a  water-bath  kept  about  45°  C.  until  it  assumes  a  deep  purple 
colour,  which  it  does  in  a  few  minutes.  Make  sections — vertical  and  horizontal — and  mount 
them  in  glycerine. 

{c)  Reduction  with  Formic  Acid. —  Place  the  cornea  in  the  juice  of  a  fresh  lemon  for  five 
minutes.  Wash  it  well,  and  transfer  it  10  a  one  per  cent,  gold  chloride  solution  for  half  an 
hour.  Wash  it  again,  and  place  it  in  a  twenty-five  per  cent,  solution  of  formic  acid  for 
twenty-four  hours.  It  must  be  kept  in  the  dark  until  the  reduction  is  complete.  This  process 
removes  all  the  epithelium,  and  is,  therefore,  not  suited  for  showing  the  terminations  of  the 
nerve-fibrils  between  the  cells  of  the  anterior  epithelium,  but  it  shows  the  nerve-fibrils  and 
corpuscles  in  the  cornea  admirably. 

Vertical  Section  of  Cornea.  EXAMINATION  (L  and  H). — Observe  the  same  general  arrange- 
ment of  parts  as  before,  but  note  the  rows   of  connective-tissue  corpuscles,  stained  of  a  deep 

P 


io6  PRACTICAL   HISTOLOGY. 

reddish  purple,  and  lying  between  the  laminae  of  the  cornea.  Trace  some  processes  communi- 
cating with  the  cells  in  the  same  plane,  and  others  with  processes  of  those  cells  that  lie  above 
and  below  them.     {Indicate  this  in  one  half  of  Y\.  XXV.,  Fig.  i.) 

Terminations  of  the  Corneal  Nerves. — The  nerve-plexus  in  the  cornea  can  be  well  studied 
in  this  section.  Observe  the  primary  nerve-plexus  in  the  corneal  tissue,  and  from  it  trace 
bundles  of  nerves  running  vertically  to  form  the  sub-epithelial  plexus.  It  is  difficult  to  find 
the  very  fine  fibrils  which  proceed  from  this  plexus  to  end  — as  an  epithelial  plexus — between 
the  cells  of  the  anterior  epithelium. 

Horizontal  Section  of  the  Cornea.  EXAMINATION  (L). — Observe  the  branched  corneal 
corpuscles  seen  on  the  flat.  They  are  deeply  stained,  so  they  stand  out  distinctly.  The 
primary  nerve-plexus  in  the  cornea  will  be  only  just  distinguishable. 

(H).  Cornea  Corpuscles. —  Note  their  flattened  characters,  their  branched  processes,  and 
their  frequent  anastomoses  with  processes  from  adjoining  cells.  Each  one  consists  of  a 
'  granular '  plate  with  a  nucleus  (PI.  XXV.,  Fig.  2). 

Nerve-Plexus. — Observe  a  meshwork  of  fine  nerve-fibrils,  with  perhaps  varicosities  on  them. 
It  may  be  that  a  bundle  of  nerve-fibrils,  constituting  an  axis-cylinder,  and  surrounded  with  a 
primitive  sheath,  may  be  found. 


THE    CELL-SPACES   OF    THE    CORNEA. 

PREPARATION. — Pith  a  frog,  and  scrape  away  the  epithelium  from  the  anterior  surface  of 
the  cornea.  Drop  into  the  eye  a  few  drops  of  a  two  per  cent,  solution  of  silver  nitrate.  Leave 
the  silver  to  act  until  the  cornea  becomes  of  a  greyish-white  colour  throughout,  which  usually 
takes  from  fifteen  to  twenty  minutes.  Excise  the  eyeball,  and  cut  out  the  cornea,  and  make 
one  or  two  cuts  in  its  margin,  and  lay  it  flat  on  a  slide  in  a  drop  of  glycerine,  and  expose  it  to 
the  light  until  it  becomes  brown  in  colour.  Similar  preparations  may  be  made  from  a  mammal, 
but  of  course  the  animal  must  be  deeply  anaesthetised  to  avoid  all  pain. 

EXAMINATION  (L  and  H). — The  cement-substance  is  stained  brown,  and  in  it  is  left  a 
series  of  clear  branched  and  anastomosing  spaces — the  cell-spaces  or  lymph-canalicular  spaces. 
They  correspond  exactly  in  shape  to  the  corneal  corpuscles,  which  partially  fill  them  (PI.  XXV., 
Fig.  3).  The  spaces  appear  to  be  empty,  because  the  cells  filling  them  are  not  affected  by  the 
silver  ;  but  if  a  section  be  stained  with  logwood  the  nuclei  of  the  corpuscles  are  brought  into 
view.  Double-staining  with  silver  and  then  gold  brings  the  corpuscles  into  view.  The  silver 
gives  the  negative  picture  and  the  gold  preparations  already  described  represent  the  positive 
picture. 

The  Fibrous  Tissue  of  the  Cornea.  PREPARATION. — Place  a  small  piece  of  the  cornea  of 
any  newly-killed  animal  in  a  solution  of  picric  acid  for  twenty-four  hours.  Tease  a  small 
piece  in  water  or  glycerine. 

EXAMINATION  (H). — Observe  the  bundles  of  wavy  fibrous  tissue  stained  yellow.  This 
preparation  need  not  be  kept. 


Plate:xx\^.  Cornea.Lens  &.  Choroid 


ly 


lylmtem  BroslitK 


loy 


THE   SCLEROTIC. 

The  sclerotic  consists  of  bundles  of  fibrous  tissue  crossing  each  other  in  all  directions  in 
the  plane  of  the  membrane.  These  bundles  become  continuous  with  the  substantia  propria 
of  the  cornea  at  its  margin.  Mixed  with  them  is  a  small  quantity  of  elastic  tissue.  Sections 
may  be  made  from  any  eye  which  has  been  hardened  in  Mi.iller's  fluid,  or  chromic  acid  and 
spirit  fluid.     Stain  the  sections  with  logwood  and  mount  them  in  dammar. 


THE    CRYSTALLINE    LENS. 

PREPARATION,  {a)  Boiling. — Boil  the  lens  of  a  fish — e.g.  cod — for  a  few  minutes,  until  it 
becomes  white.  Peel  off  the  outer  softer  part  of  the  lens,  and  tease  a  little  of  the  deeper,  less 
brittle  part,  in  Farrant's  solution. 

(d)  Glycerine  and  Nitric  Acid  Fluid.  — Place  the  eyeball  of  a  frog,  or  the  lens  of  any  other 
animal,  in  the  above  fluid  (p.  xx-xiv)  for  twenty-four  hours,  and  then  in  water  for  the  same  time. 
This  dissolves  all  -the  cement,  and  renders  the  lens-fibres  so  tough  that  they  do  not  break 
readily  when  they  are  teased.     Tease  a  small  piece  in  glycerine  as  above. 

Make  a  similar  preparation  of  the  lens  of  a  dog,  cat,  or  rabbit,  and  note  that  the  lens-fibres 
in  the  case  of  these  animals  are  not  nearly  so  markedly  serrated  (PL  XXV.,  Fig.  4). 

EXAMINATION  (H). — Observe  the  lens-fibres  of  a  codfish.  They  are  long,  narrow  bands, 
with  well-marked  teeth  or  serrations  along  their  narrow  margins,  the  teeth  of  contiguous 
fibres  dovetailing  with  one  another  (PI.  XXV.,  Fig.  5)- 

(c)  Potassic  Bichiromate. — Transverse  sections  ought  to  be  made  across  a  lens  hardened 
in  one  per  cent,  potassic  bichromate  solution  for  a  week.  Mount  them  in  Farrant's  solution. 
They  are  very  apt,  however,  to  fall  to  pieces.  The  fibres  are  hexagons  when  seen  in  transverse 
section. 

The  capsule  of  the  lens,  and  the  layer  of  epithelium  lining  it,  are  easily  obtained  from  the  eye 
of  a  frog. 

The  arrangement  of  the  lens-fibres  is  easily  made  out  in  sections  of  a  fcetal  eye. 


THE    CHOROID. 

Sections  may  be  made  through  the  coats  of  the  eyeball  of  an  ox,  hardened  in  chromic  acid 
and  spirit  mixture,  to  include  the  sclerotic  and  the  choroid.  It  consists  of  bundles  of  connec- 
tive tissue,  with  some  elastic  fibres,  many  blood-vessels  and  branched  pigment-cells.  The 
layer  of  hexagonal  pigment-cells  occurring  on  its  inner  surface  is  now  recognised  as  belonging 
to  the  retina  (PI.  XXV.,  Fig.  7). 

Pigment-Cells  of  the  Choroid.  PREPARATION. — From  the  eye  of  an  ox,  hardened  as  de- 
scribed for  the  ciliary  muscle,  remove  the  retina.  Scrape  off  a  little  of  the  dark  pigment-layer 
which  adheres  to  the  sclerotic,  and  diffuse  or  spread  it  out  in  glycerine.     .Cover. 

EXAMINATION  (H). — Observe  the  branched  flattened  cells,  like  branched  connective-tissue 


io8  PRACTICAL   HISTOLOGY. 

cells.  The  nucleus  usually  does  not  contain  pigment,  though  the  rest  of  the  cell  and  its 
processes  are  loaded  with  fine  granules  (melanin)  of  a  brown  or  black  colour  (PI.  XXV., 
Fig.  6).    . 

THE    CILIARY    MUSCLE. 

PREPARATION. —  Divide  the  eyeball  of  an  ox  or  sheep,  with  a  sharp  razor,  transversely, 
half  an  inch  behind  the  circumference  of  the  cornea,  and  remove  the  lens,  but  be  careful  to 
retain  the  choroid  and  iris.  Harden  the  anterior  half  of  the  eyeball  in  the  chromic  acid  and 
spirit  fluid  for  eight  days.  Make  horizontal  sections,  to  include  the  sclerotic,  the  cornea,  and 
the  iris.  Stain  a  section  with  picrocarmine  or  logwood  and  mount  it  in  Farrant's  solution  or 
dammar. 

EXAMINATION  (L). — Observe  the  cornea  in  the  front  part  continuous  with  the  sclerotic 
behind.  At  the  line  of  junction  notice  the  section  of  the  pigmented  iris  projecting  inwards. 
Trace  the  choroid  posteriorly,  lining  the  sclerotic.  Observe  the  fibres  of  the  ciliary  muscle 
arising  at  the  junction  of  the  sclerotic  and  cornea,  and  passing  backwards  over  and  outside 
the  choroid,  into  which  it  is  inserted.  Sections  of  the  ciliary  processes  may  be  seen.  Trace 
the  membrane  of  Descemet  of  the  cornea  backwards,  and  notice  that  at  the  junction  of  the 
cornea  and  sclerotic  it  splits  into  fine  transparent  bundles,  some  of  which  curve  round  towards 
the  iris,  whilst  others  spread  out  like  a  fan  over  the  ciliary  processes,  thus  forming  the  liga- 
mentum  pectin  at  It  111  h-idis.  At  the  junction  of  the  cornea  and  sclerotic  notice  a  small  aperture. 
This  is  a  section  of  the  venous  sinus,  or  canal  of  Sclilennn  (PI.  XXVI.,  Fig.  4). 


RETINA. 

PREPARATION.  Miiller's  Fluid  and  Spirit. — It  is  desirable  to  use  the  retinae  of  several 
animals,  because  each  shows  some  special  feature. 

(rt)  With  a  sharp  razor  divide  the  eyeball  of  a  pig,  cat,  and  ox  transversely,  and  place  the 
posterior  halves  in  a  mixture  of  three  parts  of  Miiller's  fluid  and  one  of  spirit,  and  keep  the 
preparations  in  the  dark  and  in  a  cool  place  for  a  week  or  ten  days.  Make  vertical  sections 
of  each  of  the  above  in  the  ordinary  way,  by  means  of  a  freezing  microtome,  after  the 
tissues  have  been  soaked  in  syrup  and  then  in  gum,  as  described  at  p.  xxxix.  This  is  a  most 
important  precaution  to  ensure  good  preparations.  Make  the  sections  through  the  sclerotic, 
choroid,  and  retina,  though  the  retina  is  very  apt  to  separate  from  the  other  coats.  Stain  a 
section  of  each,  with  either  logwood  or  picrocarmine.  Mount  them  in  Farrant's  solution. 
This  method  shows  the  nerve-elements  best. 

[b)  Chromic  Acid  and  Spirit  Mixture. — This  may  also  be  used,  and  shows  the  connective- 
tissue  elements  best ;  besides,  it  does  not  make  the  retina  quite  so  brittle  as  the  Muller's 
fluid. 

{c)  Osmic  Acid. — Place  the  retina  of  a  frog  and  that  of  a  fish — e.g.  cod—  in  a  few  cubic  centi- 
metres of  a  quarter  per  cent,  solution  of  osmic  acid  for  three  or  four  hours.  The  retinae  are 
rapidly  blackened  and  '  fixed.'  Soak  the  retinas  for  several  hours  in  water  to  get  rid  of  the 
surplus  osmic  acid,  and  tease  a  small  part  of  each  in  a  drop  of  glycerine,  and  cover. 

Vertical  Section  of  the  Retina  of  a  Cat  or  Pig.  EXAMINATION  (L). — Observe  from  without 
inwards :-- 


RETINA.  109 

1.  The  uvea,  ox  pigmcnt-cdls  of  the  retina,  sending  fine  processes  into  the  next  layer.  This 
layer  is  often  detached,  and  was  formerly  regarded  as  belonging  to  the  choroid. 

2.  The  layer  of  rods  and  rw/^rj  (Jacob's  membrane).  The  rods  are  easily  detected  in  groups, 
and  seen  to  be  more  numerous  than  the  cones,  which  are  much  shorter,  and  not  so  easily 
seen. 

3.  At  the  lower  end  of  the  rods  and  cones  a  well-defined  line,  indicating  the  position  of 
the  external  limiting  membrane. 

4.  The  outer  miclear  layer,  deeply  stained — red  or  blue — consisting  of  spherical  nuclei 
arranged  in  several  rows.     It  is  thicker  than  the  inner  nuclear  layer. 

5.  The  external  granular  or  internuclear  layer,  a  narrow  layer. 

6.  The  inner  nuclear  layer,  also  deeply  stained — red  or  blue — and  consisting  of  three  or 
four  rows  of  spherical  nuclei,  larger  than  those  of  the  outer  layer. 

7.  The  inner  granular  layer,  much  broader  than  the  outer  layer  of  the  same  name. 

8.  The  cellular  ox  ganglionic  layer,  consisting  of  a  single  row  of  large,  well-marked  multipolar 
nerve-cells,  each  with  a  well-defined  nucleus. 

9.  The  layer  of  optic  nerve-fibres,  merely  indicated  as  fine  threads,  as  they  have  no  white 
substance  of  Schwann. 

10.  Most  internally  is  the  internal  limiting  membra7ie.  From  this  at  regular  intervals  arise, 
by  a  broad-winged  base,  ih&  fibres  of  Micller  {cox\x\Qctive  tissue),  which  proceed  outwards  in  the 
retina  between  the  nerve-cells.  They,  may  be  traced  as  far  as  layer  number  6.  In  the  inner- 
most part  of  the  retina,  near  the  cellular  layer,  sections  of  large  blood-vessels  are  to  be  found. 
Each  of  these  parts  ought  to  be  examined  separately.  {Indicate  the  general  arrangemetit  in 
PI.  XXVI.,  Fig.  I). 

(H).  The  pigment-ceUs  or  uvea. — These  cells  are  cut  vertically,  and  are  apt  to  be  detached. 
They  send  fine  processes  into  the  next  layer. 

Rods  and  Cones. — Observe  the  rods,  more  numerous  than  the  short  club-shaped  cones. 
Each  consists  of  an  outer  and  an  inner  segment.  Notice  that  the  inner  segment  perforates 
the  outer  limiting  membrane.  Study  each  of  the  layers  in  succession.  Notice  especially  the 
nerve-cells — their  large  size,  their  well-defined  nucleus  and  nucleolus,  the  former  with  a  distinct 
envelope  around  it ;  their  processes — one  central,  which  has  a  connection  with  one  of  the  nerve- 
fibres  of  the  optic  nerve,  and  several  processes  which  proceed  outwards  and  branch  in  the 
inner  granular  layer.     Trace  the  outward  course  of  the  fibres  of  Miiller. 

The  retina  of  the  pig  is  selected  because  it  shows  so  well  the  rods  and  cones  ;  but  it  is  ad- 
visable to  study  also  a  section  of  the  retina  of  an  ox,  because  in  it  the  ganglionic  cells  are 
specially  large  and  easily  investigated. 

Retina  of  the  Frog  in  Osmic  Acid.  EXAMINATION  (H). — Tease  out  a  small  piece  in 
glycerine.  The  rods  are  very  large,  and  so  are  easily  seen.  Both  rods  and  cones  consist  of 
an  outer  and  an  inner  segment  or  member.  In  the  fresh  state  the  outer  member  is  transparent, 
bright  and  glistening,  but  it  is  rapidly  blackened  by  the  osmic  acid,  and  so  is  easily  recog- 
nised. It  not  unfrequently  shows  a  tendency  to  cleave  transversely.  Each  inner  segment  is 
pale,  and  more  or  less  granular.  They  may  be  seen  perforating  the  external  limiting  mem- 
brane, and  containing  one  of  the  nuclei  of  the  external  nuclear  layer  in  their  course.  {Indicate 
the  rods  and  these  segments  in  PI.  XXVI.,  Fig.  2.) 

The  Retina  of  a  Codfish.  EXAMINATION  (H). — This  is  specially  selected,  because  it  shows 
the  cones  so  admirably.  Select  a  cone,  and  observe  its  large  size  ;  notice  its  short  tapering 
outer  segment  and  its  large  conical  inner  one,  and  notice  how  readily  it  shows  the  transverse 


no  PRACTICAL   HISTOLOGY. 

markings  on  the  outer  segment  of  the  cone  ;  sometimes  it  may  be  found  split  transversely. 
A  view  of  the  cones  imbedded  amongst  a  great  number  of  the  narrow  rods  may  be  obtained 
from  above  (PI.  XXVI.,  Fig.  3).     Many  non-medullated  fibres  of  the  optic  nerve  will  be  found. 

DOUBLE-STAINING   OF   THE    RETINA. 

Rosein  and  Iodine-green. — Stain  a  section  of  the  retina  of  a  cat  or  ox  in  a  solution  of 
rosein  (p.  xlv),  and  then  in  iodine-green.  Mount  it  in  dammar.  The  two  nuclear  or  granular 
layers  become  of  a  greenish  colour,  and  all  the  other  parts  are  red. 

HEXAGONAL   PIGMENT-CELLS    OF   THE    RETINA. 

From  an  eye  hardened  in  chromic  acid  and  spirit  scrape  off  a  little  of  the  black  pigment 
lining  the  choroid.  A  sheet  of  flattened  hexagonal  pigment-cells  is  easily  obtained.  The 
nucleus  is  not  pigmented  (PI.  XXV.,  Fig.  7). 


THE    EYELIDS. 

The  eyelids  are  prepared  in  exactly  the  same  way  as  recommended  for  the  lips  (p.  60). 

THE    LACHRYMAL   GLANDS. 

Prepare  them   in   the  same  way  as  the  salivary  glands,  which  they  closely  resemble  in 
structure. 


Plate  XX^/i  .  Retina  k  Qliar^  Muse 


lantern.  Bi— s.lilli 


Ill 


THE  NOSE. 

(A.)     OLFACTORY   EPITHELIUM    OF   THE    FROG   OR   NEWT. 

PREPARATION,  {a)  Dilute  Alcohol.— Cut  off  the  head  of  a  newt  or  frog,  and  slit  up  the 
nostrils  with  scissors  ;  place  it  in  a  small  quantity  of  dilute  alcohol  for  twenty-four  hours. 
Stain  the  whole  head  with  picrocarmine,  and  tease  a  little  of  the  stained  epithelium  from  the 
olfactory  region  in  a  drop  of  glycerine.     This  is  the  best  method. 

\b)  Maceration  for  two  days,  in  a  quarter  per  cent,  solution  of  potassic  bichromate,  yields 
fair  results. 

{c)  Maceration  of  the  entire  head  in  a  half  per  cent,  solution  of  osmic  acid  for  five  hours, 
followed  by  steeping  in  water,  also  '  fixes '  the  cells. 

EXAMINATION  (H). — Observe  the  ordinary  columnar  epithelial  cells,  devoid  of  cilia,  but 
amongst  these  observe  the  narrow,  delicate,  spindle-shaped  cells,  each  with  a  spherical  nucleus. 
These  are  the  true  olfactot-y  cells.  To  their  free  end  a  bunch  of  fine  bristles  is  attached.  Their 
lower  or  fixed  end  consists  of  a  very  fine  filament  with  varicosities  on  it.  It  is  directly 
continuous  with  the  terminations  of  the  fibres  of  the  olfactory  nerve.  The  olfactory  cells  are 
richly  scattered  amongst  the  above-described  cylindrical  epithelium.  {Indicate  these  cells  in 
PI.  XXVI I.,  Fig.  2.) 

(B.)  Similar  preparations  may  be  made  of  the  olfactory  cells  from  the  olfactory  region  of 
the  nose  of  a  mammal.  In  this  case  the  olfactory  cells  are  devoid  of  bristles  on  their  free 
ends. 

(C.)     OLFACTORY   EPITHELIUM    OF   A  MAMMAL. 

PREPARATION. — Divide,  by  means  of  a  saw,  the  head  of  a  freshly -killed  rabbit,  dog,  guinea- 
pig,  or  sheep  in  a  longitudinal  direction,  and  parallel  to  the  nasal  septum.  Cut  out  the  nasal 
septum  so  as  to  expose  the  olfactory  region,  easily  recognised  by  its  brown  colour.  Harden 
this  in  Muller's  fluid  for  two  days,  and  then  transfer  it  to  the  chromic  acid  and  spirit 
mixture  for  a  week.  Complete  the  hardening  in  weak,  and  then  in  strong,  spirit.  Make 
vertical  sections,  and  stain  them  with  logwood,  and  mount  them  in  dammar  or  Farrant's 
solution  ;  or  a  double-staining  of  logwood  and  picrocarmine  may  be  employed. 

EXAMINATION  (L). — Observe  th'^  mucous  membrane,  covered  by  columnar  epithelium. 
The  upper  row  of  nuclei  seen  belongs  to  the  supporting  or  columnar  cells  ;  the  deeper  row 
belongs  to  the  true  olfactory  cells.  In  the  deeper  layers  of  the  epithelium,  and  also  in  the 
glands,  are  pigment-granules.  Sections  of  gland-ducts  and  alveoli,  filled  with  granular-looking 
cells — Bowman's  glands — are  readily  seen.  (H).  Examine  the  individual  parts.  {Indicate  the 
olfactory  epithelium  and  Bowman's  glands  in  PI,  XXVII.,  Fig.  i.) 


112  PRACTICAL   HISTOLOGY. 


TRANSVERSE  SECTION  OF  THE  NOSE. 

The  student  ought  to  have  a  section  through  the  whole  nasal  chamber  in  order  to  study 
the  respiratory  mucous  membrane,  nasal  septum,  the  turbinated  bones,  and  the  general 
arrangement  and  comparative  vascularity  of  the  parts. 

PREPARATION. — Inject  the  blood-vessels  of  the  head  of  a  rabbit  or  guinea-pig  with  a 
carmine-gelatine  mass  from  the  aorta.  Open  the  skull,  and  remove  the  brain  ;  disarticulate 
the  lower  jaw,  and  cut  off  the  head.  Place  it  in  Miiller's  fluid  for  three  or  four  days,  and  then 
in  the  chromic  acid  and  spirit  mi.xture  for  the  same  time.  This  fixes  the  various  elements. 
Place  it  in  the  chromic  acid  and  nitric  acid  mi.xture,  until  the  bones  are  completely  decalcified. 
Make  sections  across  the  entire  nasal  chamber  in  a  freezing  microtome.  Doubly  stain  them 
with  logwood  and  iodine-green,  and  mount  them  in  dammar.  These  form  most  instructive 
preparations. 


113 


THE   EAR. 

This  is  the  only  organ  of  which  one  cannot  give  the  student  a  section  to  carry  away 
with  him,  on  account  of  their  difficulty  of  preparation,  and  also  because  a  properly  prepared 
cochlea  yields  but  very  i&'fi  good  sections. 


THE    SEMICIRCULAR   CANALS. 

These  are  best  obtained  from  the  skate,  whose  cartilaginous  cranium  should  be  cut  away 
until  the  canals  with  their  saccule  and  utricle  are  exposed.  The  canals  are  hardened  in 
chromic  acid  and  spirit  mixture  for  a  week,  and  then  transferred  to  spirit.  The  ampullae  re- 
ceive nerves  and  may  be  hardened  in  osmic  acid.  Sections  of  these  canals  must  be  made 
with  a  freezing  microtome  after  steeping  in  syrup  and  gum. 


THE    COCHLEA. 

PREPARATION  (A). — The  guinea-pig  is  the  best  animal  to  employ.  Kill  the  animal  and 
disarticulate  the  lower  jaw,  when  the  osseous  tympanic  bulla  is  exposed.  It  lies  just  behind 
the  fossa  for  the  condyle  of  the  lower  jaw.  Cut  away  the  external  auditory  meatus,  and  re- 
move the  petrous  portion  of  the  temporal  bone  with  the  bulla  from  the  other  bones  of  the 
skull.  Open  the  bulla  with  bone-forceps,  when  a  conical  elevation — the  cochlea — is  seen. 
Remove  all  the  surrounding  bone,  so  as  to  isolate  the  cochlea.  Place  it  at  once  in  Miiller's  fluid, 
to  harden  its  delicate  tissues,  for  a  fortnight.  It  is  necessary,  however,  to  remove  the  lime- 
salts  from  the  bone,- which  is  done  by  transferring  the  organ  to  a  mixture  of  chromic  acid  and 
nitric  acid  (p.  xxxiii),  or  to  a  solution  of  picric  acid.  The  fluid  ought  to  be  shaken  from  time  to 
time,  which  greatly  facilitates  the  process  of  softening.  After  softening  preserve  it,  first  in 
weak  spirit,  and  then  transfer  it  to  strong  spirit.  When  sections  are  to  be  made  it  must  be 
steeped  in  water,  to  get  rid  of  the  spirit,  and  then  it  is  subjected  to  the  syrup-and-gum  process. 
These  substances  support  its  delicate  tissues,  and  sections  must  be  made  parallel  with  the 
modiolus,  i.e.,  across  the  turns  of  the  cochlea,  in  a  freezing  microtome.  They  must  be  manipu- 
lated with  the  greatest  care,  and  ought  to  be  stained  with  logwood  or  picrocarmine  and 
mounted  in  glycerine. 

(B).  If  the  human  cochlea  be  employed,  it  should  be  obtained  as  fresh  as  possible.  Cut 
out  the  part  of  the  temporal  bone  containing  the  cochlea,  from  the  human  subject.  Split 
this  up  in  the  axis  of  the  meatus  auditorius  internus ;  this  gives  two  pieces  of  bone,  one 
containing  the  cochlea  and  a  part  of  the  vestibule,  and  the  other  the  semicircular  canals 

Q 


114  PRACTICAL   HISTOLOGY. 

and  part  of  the  vestibule.  Place  the  pieces  at  once  in  a  mixture  of  three  parts  of  Miillcr's 
fluid  and  one  of  spirit  for  two  weeks.  After  that  transfer  them  to  a  quarter  per  cent,  solution 
of  chromic  acid  for  a  fortnight.  At  the  end  of  that  time  add  a  few  drops  of  hydrochloric  acid 
to  the  fluid  until  the  bone  is  softened.  The  bone  around  the  labyrinth  is  very  dense,  and 
requires  a  longer  time  than  the  other  parts  to  become  soft.  Wash  it  thoroughly  for  twelve 
hours  in  water,  and  place  the  whole  cochlea  for  thirty-six  hours  in  syrup  and  forty-eight  in 
gum,  and  freeze  it  in  gum  in  a  freezing  microtome. 

The  student  must  examine  a  series  of  preparations  prepared  by  the  above  methods. 

Vertical  Section  of  a  Cochlea.  EXAMINATION  (L). — Observe  the  modiolns  or  central 
column  in  the  centre,  with  the  cochlear  branches  of  the  auditory  nerve  ascending  within  it. 
Note  a  section  of  one  of  the  turns  of  the  cochlea.  Each  turn  is  divided  into  two  compartments 
by  the  lamina  spiralis.  The  upper  compartment  is  called  the  scala  vcstihnli,  and  the  lower 
one  the  scala  tympani.  Note  that  the  lamina  spiralis  consists  of  an  inner  osseous  portion  con- 
sisting of  two  plates  of  bone  between  which  runs  a  branch  of  the  cochlear  nerve  on  its  way  to 
structures  which  rest  on  the  outer  or  membranous  part  of  the  lamina.  In  the  course  of  the 
nerve  observe  the  spiral  ganglion.  The  membranous  part  consists  of  a  basilar  membrane,  on 
which  rests  the  organ  of  Corti  and  certain  epithelial  cells  which  are  connected  with  the  ter- 
minations of  the  cochlear  branch  of  the  auditory  nerve.  It  is  connected  to  the  outer  wall  of 
the  cochlea  by  the  spiral  ligament,  which  consists  of  a  mass  of  connective  tissue,  continuous 
with  the  periosteum  of  the  bone. 

Notice  a  thin  membrane  running  obliquely  upwards  and  outwards  in  the  scala  vestibuli, 
from  the  outer  part  of  the  lamina  spiral  ossea  to  the  outer  wall  of  the  cochlea  ;  this  is  the 
membrane  of  Reissner,  which  cuts  off  a  narrow  triangular  channel,  the  ductus  cochlecF,  or  scala 
media,  from  the  scala  vestibuli  proper.  The  floor  of  this  narrow  canal  is  made  up  of  the  outer  part 
of  the  osseous  lamina  and  the  basilar  membrane.  At  the  outer  extremity  of  the  osseous  lamina 
is  a  spiral  groove  excavated  like  a  C,  with  the  concavity  looking  outwards  ;  this  is  the  sulcus 
spiralis.  The  projecting  upper  edge  of  the  groove  is  called  the  crista  spiralis.  Observe  the 
organ  of  Corti,  consisting  of  an  inner  and  an  outer  set  o^  rods  resting  on  the  basilar  membrane. 
These  two  sets  of  rods  incline  towards  each  other,  their  heads  touching,  so  as  to  leave  a 
triangular  passage  or  tunnel  under  them,  which  is  filled  with  endolymph.  The  outer  rods  are 
longer  than  the  inner  ones.  Internal  to  the  inner  rods  and  external  to  the  outer  ones  are  a 
number  of  epithelial  cells,  some  of  them  with  hairs  attached  to  them — the  outer  and  inner  hair- 
cells.  A  faintly-striated  membrane  springs  from  the  upper  surface  of  the  lamina  spiralis  ossea, 
close  to  the  crista  spiralis,  and  may  be  seen  partially  to  cover  the  organ  of  Corti.  It  was 
called  by  Claudius  the  vieiiibrana  tectoria.  It  terminates  over  and  is  attached  to  the  outer 
hair-cells.  The  outer  and  inner  hair-cells  send  prolongations  through  another  membrane 
which  covers  them — the  membrana  reticularis.   {Indicate  the  general  arrangement  in  PI.  XXVII., 

Fig.  3-) 

(H).  The  membrane  of  Reissner  consists  of  a  very  delicate  hyaline-membrane  covered  on 
each  surface  with  a  single  layer  of  epithelium.  Its  surface,  directed  towards  the  cochlear  canal, 
is  composed  of  polyhedral  flattened  cells,  whilst  the  vestibular  surface  is  covered  by  very  thin 
endothelial  cells.  The  membrana  basilaris  has,  on  its  upper  surface,  many  epithelial  cells, 
some  of  them  specially  to  form  the  organ  of  Corti,  and  others  in  which  the  cochlear  branch  of 
the  auditory  nerve  probably  terminates,  and  which  are,  in  fact,  its  '  end-organs.'  The  basilar 
membrane  consists  of  one  or  two  layers  of  hyaline  membrane,  which  are  covered  on  the  tym- 
panic side  by  a  single  layer  of  endothelial  cells.     On  the  upper  or  vestibular  surface  of  the 


Plate XXVn .  Nose  &  Cochlea 


Twfirvtem.  Bros.litK 


COCHLEA.  1 1 5 

membrane  rests  Corti's  organ.  External  and  internal  to  it  lie  several  rows  of  cells.  Beginning 
at  the  outer  end  of  the  basilar  membrane  observe  the  supporting  cells  of  Hcnsen,  and  then  a 
row  of  cells  with  fine  bristles  projecting  from  their  upper  surface — the  outer  hair-cells.  These 
cells  occur  in  three  or  four  rows.  Passing  onwards  we  come  to  the  arch  of  Corti,  composed  of 
an  outer  and  an  inner  rod  or  pillar,  each  in  a  single  row.  Each  rod  has  the  shape  of  an  /,  and 
they  are  so  arranged  as  to  touch  with  their  upper  part  or  head,  whilst  the  base  rests  on  the 
membrana  basilaris,  thus  leaving  a  triangular  space  between  them.  Each  rod  consists  of  a 
head,  a  narrow  middle  part  or  body,  and  a  broad,  wing-shaped  base  or  foot.  The  head  of 
the  inner  rod  has  a  concavity  on  its  outer  surface  which  receives  the  convex  surface  of  the 
outer  rod.  A  flat  process  or  head-plate  proceeds  outwards  which  helps  to  cover  in  the  head 
of  the  outer  rod.  It  terminates  in  the  membrana  or  lamina  reticularis.  The  outer  rod  also 
sends  outwards  a  short  process  which  forms  the  first  phalanx  of  the  membrana  reticularis. 
The  outer  rods  are  longer,  thicker,  and  more  numerous  than  the  inner  rods.  At  the  foot  of 
each  rod,  on  the  surface  directed  towards  the  triangular  tunnel  under  the  rods,  is  a  small 
nucleated  mass  of  protoplasm. 

The  inner  hair-cells  of  Deiters  lie  immediately  inside  the  inner  rods.  They  occur  in  a 
single  row,  and  possess  fine  hairs,  projecting  from  their  free  surface  just  like  the  outer  hair-cells. 
Internal  to  these  there  is  a  row  of  nucleated  columnar  epithelial  cells,  which  support  these 
cells.  They  are  continued  inwards  over  the  membranous  portion  of  the  membrana  basilaris 
as  far  as  the  sulcus  spiralis.  The  lamina  or  meuibrana  reticularis  will  be  seen  in  section.  It 
is  a  cuticular,  hyaline,  very  resistant  membrane  which  stretches  outwards  from  the  heads  of 
Corti's  rods  to  the  outer  hair-cells.  In  it  exist  a  number  of  round  or  oval  apertures,  through 
which  the  outer  hair-cells  project.  The  parts  of  this  membrane  which  exist  between  these 
holes,  being  shaped  like  a  finger,  were  called  phalanges  by  Deiters.  {Indicate  the  menibrane  of 
Reissner,  Corti s  rods  and  cells,  and  the  membrana  tectoria  in  PI.  XXVII. ,  Fig.  4.) 

The  organs  of  taste  (p.  62)  and  touch  (p.  95)  have  already  been  alluded  to. 


Q2 


ii6  PRACTICAL   HISTOLOGY. 


THE  MALE   GENERATIVE   ORGANS. 


THE    TESTIS. 

PREPARATION,  {a)  Chromic  acid  and  Spirit  Mixture. — Use  the  testis  of  a  dog,  rabbit,  or 
guinea-pig.  Make  several  cuts  across  the  testis  with  a  sharp  razor,  taking  care  to  squeeze  the 
organ  as  little  as  possible,  and  place  it  in  the  above  fluid.  Change  the  fluid  frequently,  and 
at  the  end  of  ten  days  transfer  it  to  weak,  and  then  to  strong,  spirit.  Make  transverse 
sections — by  freezing — through  the  whole  organ  ;  stain  them  with  logwood,  and  mount 
them  in  dammar.  They  are  apt  to  fall  to  pieces,  but  by  floating  the  section  in  water  all 
difficulty  is  overcome. 

{J})  Miiller's  fluid. — Harden  the  testis  for  three  weeks  in  Miiller's  fluid,  and  then  in  spirit. 
The  testis  is  rendered  very  tough  by  this  method,  and  it  is  less  apt  to  fall  to  pieces. 

{c)  Interstitial  Injection  of  Osmic  Acid. — Inject  into  the  testis,  with  a  subcutaneous  syringe, 
a  one  per  cent,  solution  of  osmic  acid,  and  harden  the  organ  in  alcohol.  This  method  shows 
the  intertubular  connective  tissue  very  well  after  being  stained  with  logwood. 

Transverse  Section  of  the  Testis.  EXAMINATION  (L). — Observe  the  capsule,  consisting  of 
an  outer  layer — the  tunica  adnata,  or  the  visceral  layer  of  the  tunica  vaginalis — and  an  inner, 
dense,  fibrous  coat — the  tunica  albuginea.  Between  these  two  layers  oval  openings  are  seen  : 
they  are  sections  of  lymphatic  vessels.  From  the  capsule  septa  radiate  inwards  towards  the 
corpus  Highmori,  so  as  to  subdivide  the  organ  into  a  series  of  compartments,  each  one  of 
which  is  filled  with  small  convoluted  tubes — the  tubuli  seuiiniferi — which,  of  course,  are  cut 
in  every  possible  direction.  These  compartments,  when  filled  with  the  tubules,  constitute  the 
lobules  of  the  testis.  These  septa  have  a  decidedly  lamellar  structure,  and  they  carry  the 
large  blood-  and  lymph-vessels.  Note  these  tubes  of  nearly  uniform  diameter,  and  observe  the 
small  amount  of  intertubular  connective  tissue,  which  takes  the  form  of  lamellae  of  connective 
tissue  covered  with  endothelial  cells.  The  interlamellar  spaces  communicate  freely  with  each 
other,  and  form  the  origin  of  the  lymphatics  of  the  testis  (p.  Ii8). 

In  some  animals  (dog,  cat,  and  boar),  columns  of  peculiar  granular,  nucleated  cells  are 
found  between  the  tubules.  In  some  animals  (guinea-pig)  they  are  pigmented,  but  their  exact 
nature  is  unknown. 

The  tubuli  seminiferi  are  relatively  large,  wavy,  and  convoluted  tubes  of  uniform  diameter, 
and  are  cut  in  every  direction.  Each  tube  is  seen  to  consist  of  a  thick  membrana  propria,  of 
a  clear  hyaline  character,  with  flattened  nuclei  in  it,  so  that  it  is  probably  endothelial  in  its 
nature.  It  is  more  or  less  completely  filled  with  seminal  cells.  It  may  be  possible  to  trace  the 
tubules  in  their  course  towards  the  corpus  Highmori,  where  they  empty  themselves  into  the  vasa 


TESTIS. 


117 


recta,  which  in  the  corpus  Highmori  form  the  rete  testis.  These  tubes  are  much  thinner  than 
the  seminal  tubes,  and  there  is  a  constriction  where  the  one  tube  passes  into  the  other.  They 
are  Uned  by  a  single  layer  of  low,  columnar  cells.  They  lead  into  the  wider  coni  vasculosi  and 
the  epididymis  (p.  117).     (^Indicate  tlie  capsule  and  sections  of  the  tubules  in  PI.  XXVIII.,  Fig.  i.) 

(H).  Study  the  tubuli  seminiferi,  especially  in  a  transverse  section.  Observe  the  thick 
membrana  propria  ;  within  it  are  the  seininal  cells,  arranged  in  several  layers.  The  outer 
seminal  cells,  next  the  membrana  propria,  consist  of  a  single  layer  of  transparent,  polyhedral, 
nucleated,  and  faintly-defined  cells.  Internal  to  these  are  several  layers  of  cells — the  inner 
seminal  cells.  They  are  polyhedral,  nucleated,  somewhat  granular  cells,  though  they  are 
usually  more  spherical  in  shape  next  the  lumen  of  the  tube. 

Study  the  innermost  cells  : — some  of  them  may  be  found  with  their  nucleus  partially 
divided,  or  even  divided  into  two  daughter  nuclei,  and  then  the  whole  cell  divides,  so  as  to 
give  origin  to  two  dajighter  cells.  These  cells  undergo  further  changes,  and  give  origin  to 
the  spermatozoa,  and  are  hence  called  spermatoblasts.  These  cells,  after  certain  intermediate 
changes  in  their  nuclei  and  cell-contents,  split  up  into  a  group  of  young  spermatozoa,  which 
are  arranged  in  fan-shaped  groups,  the  head  or  nucleus  of  each  spermatozoon  being  directed 
towards  the  membrana  propria  of  the  tube.  This  development  of  the  spermatozoa  in  groups 
is  quite  characteristic,  and  several  groups  may  be  seen  in  a  single  tube  ;  afterwards  they 
become  detached  to  constitute  spermatozoa.  They  are  best  seen  in  dammar  preparations.  It 
is  to  be  remembered  that  all  these  stages  are  not  found  in  a  section  of  any  one  tube,  but 
several  tubes  must  be  examined  to  see  all  the  stages.  {Indicate  the  cells  lining  a  tubule  and 
the  developing  spermatozoa  in  PI.  XXVIII. ,  Fig.  2.) 


THE    EPIDIDYMIS   AND   VAS    DEFERENS. 

PREPARATION. —  Exactly  the  same  as  directed  for  the  testis.  It  is  convenient  to  use  the 
epididymis  of  a  sheep.  Make  transverse  sections,  stain  them  in  logwood,  and  mount  them  in 
dammar,  and  do  the  same  with  the  vas  deferens. 

Transverse  Section  of  the  Epididymis.  EXAMINATION  (L  and  H).— We  traced  the  seminal 
tubules  until  they  formed  the  rete  testis.  This  system  of  tubes  opens  into  ten  or  twenty  vasa 
efferentia,  which  form  the  coni  z'asculosi,  which  again  open  into  the  canal  of  the  epididymis,  and 
this  again  into  the  vas  deferens.  Sections  of  the  vasa  efferentia  and  the  convoluted  tube 
of  the  epididymis  are  well  seen,  the  latter  being  much  larger  than  the  former.  Each  transverse 
section  of  the  epididymis  shows  tubes  with  a  relatively  large  lumen,  lined  with  columnar 
epithelium,  which  rests  on  a  membrana  propria,  supported  by  non-striped  muscle.  The 
epithelium  is  columnar  and  ciliated,  and  the  cilia  are  relatively  long.  {Indicate  the  general 
arrangement  in  PI.  XXVIII.,  Fig.  3,  and  the  ciliated  epithelium  in  Fig.  4.) 

At  the  bases 'of  these  cells  branched  pigment-cells  are  found,  especially  in  the  testis  of  the 
sheep.  Notice  the  normal  amount  of  interstitial  connective  tissue,  in  which  nerves  and  blood- 
vessels may  be  recognised. 

The  vas  deferens  has  a  mucous,  a  muscular,  and  an  outer  coat,  or  adventitia.  The  epithe- 
lium of  this  canal  is  columnar,  and  carries  cilia  only  in  that  part  next  the  epididymis. 

The  blood-vessels  of  the  testis  are  studied  in  the  ordinary  way. 


]i8  PRACTICAL   HISTOLOGY. 


THE    LYMPHATICS   OF    THE    TESTIS. 

PREPARATION. — Thrust  the  nozzle  of  a  syringe  into  a  perfectly  fresh  testicle,  and  inject 
into  its  substance  a  two  per  cent,  watery  solution  of  Berlin-blue.  The  lymphatics  in  the  tunica 
albuginea  and  spermatic  cord  are  rapidly  filled.  Harden  the  testis  in  alcohol,  and  make  sec- 
tions, which  are  to  be  stained  with  picrocarmine  and  mounted  in  dammar,  or  in  Farrant's  solu- 
tion. The  blue  colouring  matter  passes  into  the  origin  of  the  lymphatics  between  the  tubes, 
but  none  of  it  passes  into  them. 

EXAMINATION  (L  and  H). — Observe  the  seminal  tubules,  stained  of  a  yellowish-red 
colour,  and  between  them  note  the  distribution  of  the  blue  colouring  matter,  which  has  passed 
into  the  lymphatics. 

THE    SPERMATOZOA. 

(A.)     IN   THE    LIVING  CONDITION. 

1.  PREPARATION  in  the  Newt. — Kill  a  large  male  newt  (Triton  cristatus),  which  may  be 
readily  recognised  by  the  serrated  crest  along  its  back.  Remove  the  viscera,  wlien  the  testes 
— two  or  three  on  each  side— will  be  found  as  small  round  bodies,  lying  close  to  the  spinal 
column.  Remove  one  of  these,  make  a  cut  into  it,  and  press  out  a  little  of  its  milky  contents 
on  to  a  slide,  add  a  drop  of  salt  solution  ;  cover  and  examine. 

EXAMINATION  (H). — Observe  the  seething  mass  of  spermatozoa.  Each  one  consists  of 
a  so-called  licad,  to  which  is  attached  a  long,  curved  prolongation,  the  tail  ox  filament.  Study 
one,  and  note  that  the  tail  moves  with  a  rapid  lashing  motion,  not  unlike  the  action  of  a 
cilium,  with  the  result  of  propelling  onwards  the  spermatozoon. 

It  is  well  to  examine  these  spermatozoa  with  a  higher  power,  with  a  lens  of  at  least  ^-inch 
focal  distance.  Each  spermatozoon  is  then  seen  to  consist  of  a  head,  and  next  to  it  is  the 
middle  piece  of  Schweigger-Seidel,  and  attached  to  this  is  the  long,  pointed  filament  or  tail. 
A  long  spiral  filament  is  attached  to  this  by  a  very  transparent  membrane  (Leydig,  H. 
Gibbes). 

2.  In  a  Mammal,  e.g.  a  Sheep. — Make  a  cut  into  the  globus  major  of  a  perfectly  fresh  testicle, 
and  press  out  a  little  of  the  milky  seminal  fluid,  and  examine  it  as  above. 

(B.)     PERMANENT   PREPARATIONS. 

1.  Spermatozoa  of  the  Newt, — Harden  the  testis  of  a  newt  in  five  per  cent,  ammonium 
chromate  for  twenty-four  hours  ;  wash  away  all  the  colouring  matter,  and  divide  a  testis,  and 
then  press  out  a  little  of  the  fluid,  which  is  next  mixed  with  a  drop  of  glycerine,  covered  and 
preserved  in  the  ordinary  way.  The  spermatozoa  of  a  newt  may  be  stained  with  logwood  and 
an  aniline  dye,  but  the  process  is  rather  difficult,  and  requires  much  time  and  practice. 

2.  Mammalian  Spermatozoa,  ia) — Place  a  little  glycerine  in  a  watch-glass,  and  add  to  it  a 
few  drops  of  absolute  alcohol.  Make  a  cut  into  the  globus  major  of  a  perfectly  fresh  testis. 
Press  out  some  of  the  fluid  into  the  watch-glass,  and  mix  it  thoroughly  with  the  glycerine 
fluid.     Place  a  drop  of  this  on  a  slide,  and  cover  (PL  XXVIH.,  Fig.  5). 

{b)  Another  way  is  to  place  a  drop  of  seminal  fluid  on  a  slide  and  allow  it  to  dry,  and  then 
cover  and  seal  it  up  diy,  without  the  addition  of  any  fluid. 


PL^EJCXVm  Testis  &  Ovary 


MinLem  Bi'os  lith 


119 


THE    PROSTATE    GLAND   AND    PENIS. 

These  are  best  hardened  in  the  chromic  acid  and  spirit  mixture.  It  is  well  to  examine  a 
section  of  a  penis  with  its  blood-vessels  injected,  to  study  the  venous  sinuses  in  the  corpus 
cavernositm  penis.  The  glans  penis,  when  stained  with  gold  chloride  reduced  by  the  acetic 
acid  method,  shows  the  mode  of  termination  of  medullated  nerve-fibres  between  the  epithelial 
cells  covering  it,  and  also  their  mode  of  termination  in  end-bulbs.  In  the  prostate  numerous 
sections  of  branched  tubular  glands  lined  with  columnar  epithelium,  and  supported  by  much 
non-striped  muscle,  are  to  be  found. 


I20  PRACTICAL   HISTOLOGY. 


THE  FEMALE   GENERATIVE   ORGANS. 


THE    OVARY. 

PREPARATION,  [a)  Chromic  Acid  and  Spirit  Mixture. — Harden  the  entire  ovary  of  a  cat, 
rabbit,  or  bitch  in  the  above  fluid  for  seven  to  ten  days,  and  then  transfer  it  to  weak,  and 
afterwards  to  strong,  spirit.  Make  transverse  sections,  stain  some  with  logwood,  and  mount  one 
in  dammar,  and  another  in  Farrant's  solution. 

{b)  Mttller's  Fluid. — Harden  the  entire  ovaries  of  any  animal  in  Miiller's  fluid.  Change  the 
fluid  at  the  end  of  the  first  day,  and  continue  the  hardening  for  three  weeks.  The  specimens 
must  on  all  occasions  be  handled  very  gently,  to  avoid  the  removal  of  the  germinal  epithelium 
covering  them.  Transfer  them  to  weak,  and  then  to  strong  spirit.  Both  methods  yield  ex- 
cellent results. 

Transverse  Section  of  the  Ovary.  EXAMINATION  (L). — Observe  the  framework,  which  con- 
sists of  (i)  the  tissue  of  the  hilum,  and  (2)  the  stroma  of  the  parenchyma.  The  tissue  of  the 
hilum  consists  of  loose  areolar  tissue,  with  many  large  blood-vessels.  The  walls  of  the  arteries 
are  easily  made  out,  because  they  are  very  thick.  The  stroma  consists  of  bundles  of  clear 
fusiform  nucleated  cells,  with  a  small  amount  of  fibrous  tissue.  It  is  doubtful  whether  these 
fusiform  cells  are  muscular  or  not.  Near  the  surface  of  the  ovary  are  one  or  two  layers  of 
these  cells,  which  usually  contain  no  ova.  In  the  stroma  small  groups  of  polyhedral  cells — 
interstitial  cells — the  rudiments  of  the  Wolffian  bodies,  like  those  occurring  in  the  stroma  of 
the  testis,  may  be  found.  Note  the  single  layer  of  short  columnar  epithelial  cells — or  germinal 
epitJielium  of  Waldeyer — covering  the  surface  of  the  ovary. 

Note  the  Gi-aafian  follicles  lying  in  the  stroma,  and  observe  their  distribution.  They  are 
of  various  sizes  and  shapes,  and  the  larger  ones,  i.e.  the  more  developed,  lie  in  the  deeper  part 
of  the  stroma,  whilst  the  smaller  ones  form  a  layer  two  or  three  deep  in  the  superficial  part  of 
the  ovary.     {Indicate  the  distribution  of  the  follicles  in  PI.  XXVIII.,  Fig  6.) 

Study  a  large  Graafian  Follicle  (L). — Observe  the  spindle-cells  arranged  more  or  less  con- 
centrically around  it  to  form  the  tunica  fibrosa.  Within  this  a  membrana  propria,  lined  with 
several  layers  of  cells,  which  constitute  the  membrana  granulosa  of  the  follicle,  and  placed 
more  or  less  excentrically  is  the  ovum.  The  ovum  is  imbedded  in,  and  rests  on,  some  of  the 
cells  of  the  membrana  granulosa,  and  those  cells  on  which  it  rests  form  the  cumubis  proligerus, 
and  these  cells  are  continued  around  the  ovum,  as  the  tunica  granulosa,  so  as  to  embrace  it. 
A  cavity,  which  is  filled  with  an  albuminous  fluid,  the  liquor  folliciili,  exists  between  these  two 
layers  of  the  membrana  granulosa.  The  ovum  is  a  complete  cell,  and  consists  externally  of  a 
hyaline  cell-membrane,  the  zona  pellucida  ;  within  this  is  the  transparent  yolk  or  vitellits.  Im- 
bedded usually  in  one  side  of  the  yolk  is  the  germinal  vesicle  (nucleus),  with  its  included 
germinal  spot.     {Indicate  the  ovum  and  membrana  granulosa  in  PI.  XXIX.,  Fig.  i.) 


PI^TEXXK  .  OvARXEaLLOPIAI^  TUBEjiTERUS^ 


JXCCCl 


&     (o 


e^ 


(i\ 


7\  i 

H 


Mmteni.  Bros.litK 


OVARY.  121 

(H).  Study  each  of  these  parts  from  within  outwards.  In  the  vitellus,  yolk-granules  and 
a  fine  reticulum  of  fibres  may  be  detected. 

Study  a  Small,  Unripe,  Graafian  Follicle.  (H). — These  are  found' near  the  surface  of  the 
section.  Notice  the  ovum,  with  a  relatively  thin  zona  pellucida,  and  the  membrana  granulosa, 
consisting  of,  perhaps,  only  two  layers  of  cells,  and  not  yet  separated  into  two  distinct  layers 
by  the  appearance  of  the  liquor  folliculi  between  them.  All  gradations  are  found  between 
these  and  a  follicle  ready  to  burst  and  discharge  its  ovum.  {Indicate  tlicsc,  and  the  epithelium 
covering  the  ovary,  in  PI.  XXIX.,  Fig.  2.) 


THE    DEVELOPMENT   OE   GRAAEIAN    FOLLICLES   AND   OVA. 

The  -ovaries  of  fcetal  kittens  or  rabbits  are  prepared  in  the  same  way  as  directed  for  the 
adult  ovar>'  (p.  1 20).  Sections  show  the  development  of  the  ova  from  the  germinal  epithelium 
covering  the  ovary. 

CORPORA   LUTEA. 

These  are  easily  studied  in  the  ovary  of  a  cow,  where  they  are  very  large.  Small  pieces  of 
one  of  them  ought  to  be  macerated  in  very  dilute  potassic  bichromate,  to  isolate  the  large 
branched  and  pigmented  cells  which  compose  them.  Coloured  crystals  of  hsematoidin  may  be 
found.  Other  pieces  ought  to  be  hardened  in  Miiller's  fluid  for  two  weeks,  and  sections  made 
to  study  the  arrangement  of  the  cells  which  compose  them. 


THE    FALLOPIAN    TUBE. 

PREPARATION.— Cut  the  Fallopian  tube  of  a  cat  or  bitch  into  pieces  an  inch  long,  and 
harden  them  for  four  days  in  the  chromic  acid  and  spirit  mixture.  Make  transverse  sections, 
and  stain  them  with  logwood,  and  mount  them  in  dammar. 

EXAMINATION  (L  and  H).  Observe  the  serous,  or  outer  coat,  composed  of  fibrous  tissue, 
and  within  this  the  muscuhir  coat,  composed  externally  of  a  few  muscular  fibres,  arranged  longi- 
tudinally, and  therefore  cut  transversely,  whilst  within  these  is  a  much  thicker  circular  coat. 
Then  follows  the  mucous  coat,  consisting  of  fibrous  tissue,  containing  many  blood-vessels. 
It  is  thrown  into  longitudinal  folds  by  the  contraction  of  the  circular  muscular  fibres,  and  is 
lined  by  columnar  ciliated  epithelium.  It  contains  no  glands.  Do  not  mistake  the  depressions 
between  the  folds  of  the  mucous  membrane  for  glands.  {Indicate  part  of  tlie  wall  of  the  tube  in 
PI.  XXIX.,  Fig.  SO 


THE    UTERUS. 

PREPARATION.— Remove  the  uterus  from  a  cat  or  bitch — preferably  from  an  animal 
which  has  borne  young — cut  it  in  pieces  an  inch  long,  and  harden  them  for  five  days  in  chromic 
acid  and  spirit  mixture,  and  then  transfer  them  to  spirit.  Make  transverse  sections,  and  stain 
them  with  logwood,  and  mou;it  them  in  dammar. 

R 


122  PRACTICAL   HISTOLOGY. 

EXAMINATION  (L). — Observe  («)  the  serous  coat,  {b)  the  muscular  coat,  very  thick,  and 
composed  of  bundles  of  non-striped  muscle,  arranged  in  many  layers.  The  mucous  coat  con- 
sists of  fibrous  tissue,  and  is  lined  by  columnar  ciliated  epithelium  (the  cilia  are  difficult  to 
preserve),  and  in  it  are  seen  sections  of  the  uteruie  glands,  v/hxch  are  cut  in  every  direction,  but 
they  are  simple  tubular  glands,  often  branched  at  their  lower  extremities,  and  they  are  lined 
throughout  by  ciliated  epithelium  (PL  XXIX.,  Fig.  3). 

(H).  Observe  specially  the  uterine  glands,  with  their  membrana  propria  lined  by  ciliated 
columnar  cells.     {Indicate  a  gland  in  PI.  XXIX.,  Fig.  4.) 

Similar  preparations  ought  to  be  made  of  the  cervix  uteri,  e.g.,  of  a  cow. 


THE    MAMMARY   GLAND. 

PREPAKATION. — The  best  specimens  are  obtained  from  a  gland  taken  from  an  animal 
near  the  full  period  of  gestation.  Harden  small  pieces  for  a  week  in  the  chromic  acid  and 
spirit  mixture  ;  make  sections,  stain  them  with  logwood,  and  mount  them  in  dammar. 

EXAMINATION  (L). — Observe  the.  framework,  which  consists  of  septa,  chiefly  of  lamellar 
connective  tissue,  which  subdivide  the  gland  into  a  series  of  small  polygonal  lobules.  In  these 
septa  sections  of  the  large  lactiferous  or  milk  ducts,  and  large  blood-vessels  are  found. 

Study  a  lobule  (L  and  H). — Observe  its  shape,  and  note  that  it  consists  of  a  large  number 
oi gland  alveoli,  supported  by  a  very  small  amount  of  connective  tissue. 

Study  an  alveolus  (H). —  Observe  its  membrana  propria,  lined  by  a  single  layer  of  short 
columnar  epithelial  cells.  Sections  of  the  large  lobular  ducts  will  be  readily  met  with.  Fat- 
globules,  i.e.  milk-granules,  may  be  seen  within  the  protoplasm  of  these  cells.  Osmic  acid 
readily  reveals  the  presence  of  fatty  particles  in  the  cells  by  blackening  them.  {Indicate  the 
alveoli,  with  the  cells  lining  them,  in  PL  XXX.,  Fig.  i.) 


THE    MILK. 

Place  a  drop  of  fresh  milk  on  a  slide,  apply  a  cover-glass  and  examine  (H).  It  consists  of 
a  large  number  of  oil-granules,  of  various  sizes,  floating  in  a  fluid.  Each  globule  consists  of 
an  albuminous  envelope  (Ascherson's  membrane),  enclosing  a  globule  of  oil.  Add  acetic  acid, 
which  partially  dissolves  the  envelope,  and  then  the  oil-globules  run  into  little  heaps  or  clusters. 
Osmic  acid  blackens  them.  Do  not  preserve  them.  {Delineate  in  the  upper  half  of  PL  XXX., 
Fig.  2,  ordinary  milk,  and  in  the  lower  half  the  effect  of  acetic  acid  on  it.) 

In  the  milk  shortly  after  delivery  many  large  nucleated  cells,  filled  with  milk-globules — 
the  colostrum  corpuscles — are  to  be  found.     Examine  these  (H). 


THE    PLACENTA. 

PLACENTA   OF   A    CAT   OR   GUINEA-PIG. 

PREPARATION. — The  animal   ought   to   be  killed   about  the  middle  of  the    period  of 
gestation.     Open  the  abdomen  and  expose  the  uterus.     The  uterus   is  cautiously  opened, 


PlateXXX,  Mamma.MilkPlacenta. 


Mintem  Bro  B  itli 


PLACENTA.  123 

and  care  is  taken  not  to  sever  the  attachment  of  the  placenta  to  the  uterus.  Harden  the 
uterus,  placenta,  and  attached  foetus,  which  is  still  within  the  amniotic  cavity,  in  the  chromic 
acid  and  spirit  mixture  for  two  weeks,  and  then  transfer  them  to  dilute  spirit,  and  afterwards 
to  strong  spirit.  The  foetus  can  be  used  for  making  sections  of  various  organs.  Make  vertical 
sections  through  the  uterus  and  the  placenta,  and  stain  them  with  logwood  or  picrocarmine, 
and  mount  them  in  dammar  or  Farrant's  solution. 

EXAMINATION  (L). — Observe  the  uterus,  with  the  placenta  lying  internal,  and  firmly 
attached  to  it.  The  placenta  consists  of  a  narrow  maternal  part  next  the  uterus,  and  a  much 
thicker  part — the  fcetal  portion.  In  the  uterus  numerous  sections  of  the  enlarged  uterine 
sinuses  are  seen.  The  foetal  portion  consists  of  richly  branched  villi,  which  are  separated 
from  each  other  by  a  relatively  small  amount  of  delicate  connective  tissue,  and  a  layer  of  cells, 
which  covers  each  villus.  Within  these  villi  note  the  loops  of  blood-vessels.  Their  distribu- 
tion is  easily  seen,  becau.se  they  still  contain  the  coloured  blood-corpuscles. 

FRESH    HUMAN    PLACENTA. 
PREPARATION. — Tease  a  small  piece  of  a  perfectly  fresh  placenta  in  salt  solution. 

EXAMINATION  (L). — Observe  the  large  number  of  isolated  and  richly  branched  placental 
villi.  Each  one  contains  a  blood-vessel,  whose  course  is  easily  seen,  because  it  is  still  filled 
with  blood-corpuscles.  A  capillary  loop  is  sent  into  each  of  the  secondary  branches  on  the 
villus.  The  villus  is  covered  by  a  layer  of  epithelium,  whose  nuclei  are  easily  revealed  by 
staining  the  section  with  magenta  solution.  This  epithelial  covering  varies  in  thickness  on 
the  same  villus.  A  delicate  mucous  tissue  exists  around  the  blood-vessels.  {^Indicate  the  shape 
and  general  characters  of  the  villi  in  PL  XXX.,  Fig.  3.) 

(H).  Observe  the  capillary  blood-vessels  filled  with  blood-corpuscles,  and  between  the 
vessels  note  the  corpuscles  of  the  mucous  tissue  which  supports  them.  Study  the  epithelial 
covering.  It  is  low,  columnar  epithelium,  with  here  and  there  masses  of  protoplasm,  which 
contain  many  nuclei.     (^Indicate  the  epithelial  covering  and  blood-vessels  in  PL  XXX.,  Fig.  4.) 

HOW   TO    PRESERVE   THE    HUMAN    PLACENTA. 

(1)  Osmic  Acid. — A  small  piece  of  the  fresh  placenta  ma}-  be  teased  in  a  one  per  cent, 
solution  of  osmic  acid,  and  mounted  in  glycerine. 

(2)  Dilute  AIcohoL — Maceration  for  forty-eight  hours  in  this  fluid  is  an  excellent  means  of 
isolating  the  villi.     They  may  be  stained  with  picrocarmine. 

(3)  Miiller's  Fluid. — Harden  very  small  pieces  of  a  perfectly  fresh  placenta  for  three  weeks 
in  this  fluid.  Make  vertical  sections,  and  treat  them  as  directed  for  the  placenta  of  a 
guinea-pig. 


R  2 


124  PRACTICAL    HISTOLOGY. 


THE   FCETAL    MEMBRANES. 


THE    DECIDU^. 

The  decidua  vera  and  reflexa  can  be  separated  from  each  other  near  the  placenta. 
Remove  a  little  of  the  tissue  from  the  outer  surface  of  the  decidua  reflexa,  and  examine  it  in 
salt  solution.  It  will  be  found  to  be  nearly  covered  with  fusiform  cells,  which  are,  for  the 
most  part,  filled  with  fatty  particles.  These  cells  may  be  stained  with  methyl-aniline,  and 
preserved  in  a  saturated  solution  of  acetate  of  potash. 

The  substance  of  the  decidua  consists  of  a  tissue  closely  resembling  mucous  tissue.  If  a 
small  piece  be  treated  with  gold  chloride  (p.  xlv)  branched  connective-tissue  corpuscles  are 
obtained. 


THE   AMNION. 

K  small  piece  of  this  membrane,  after  being  hardened  for  five  days  in  Miiller's  fluid,  is 
thoroughly  washed,  and  placed  on  a  slide,  and  stained  with  picrocarmine.  Observe  on  its 
inner  surface  a  layer  of  somewhat  flattened  cells. 


THE    UMBH^ICAL   CORD. 

PREPARATION. —  Cut  a  fresh  cord  into  pieces  an  inch  in  length,  and  harden  them  in 
Miiller's  fluid  for  two  days,  and  afterwards  in  the  chromic  acid  and  spirit  mixture  for  a  week. 
Make  transverse  sections.  Stain  one  with  logwood,  and  mount  it  in  dammar,  and  another 
with  picrocarmine,  and  mount  it  in  Farrant's  solution. 

EXAMINATION  (L). — Observe  the  circular  shape  of  the  cord,  containing  sections  of  three 
blood-vessels,  £e.  two  umbilical  arteries  and  a  larger  umbilical  vein.  These  are  surrounded  by 
a  modified  mucous  tissue  (p.  28),  called  Wharton^s  jelly,  and  outside  all  is  a  layer  of  thin, 
nucleated,  polygonal  cells.  The  tissue  is  denser  around  the  blood-vessels.  In  Wharton's  jelly 
are  found  spaces — the  sections  of  canals  that  run  in  the  cord.  They  are  most  obvious  near 
the  circumference.  Note  the  mucous  corpuscles  amongst  the  fibres.  {^Indicate  the  uinbilical 
arteries  and  vein,  and  Wharton's  jelly,  in  PI.  XXX.,  Fig.  5.) 

(H).  Examine  the  walls  of  the  arteries  and  vein,  and  note  their  great  thickness.  Study 
the  branched  corpuscles  of  Wharton's  jelly.  {Indicate  the  spaces  and  the  mucous  tissue  in  PI. 
XXX.,  Fig.  6.) 


INDEX. 


o^Jo 


ACE 

ACETIC  acid,  xx 
Adenoid  tissue,  29 
Adipose  tissue,  27 

development  of,  27 

Adjustment,  coarse  and  fine,  xxii 

—  of  objective,  xxii 
Air-cells  of  the  lung,  56 
Alcohol,  absolute,  xxxi 

—  dilute,  xxxiv 

—  methylated,  xxxii 

—  rectified,  xxxiv 
Ammonium  bichromate,  xxxii 

—  chromate,  xxxii 

—  picrocarminate,  xliii 
Amnion,  124 
Amosboid  movement,  4 
Aniline  blue  for  peptic  cells,  67 
Aniline  blue-black  for  nerve-tissues,  xliv 
Aniline  dyes,  xliv 

Aniline  violet,  xliv 

Anterior  roots,  97 

Aorta,  50 

Apparatus  necessary  for  histological  work,  xix 

Appendix  vermiformis,  70 

Aqueous  humour,  xxx 

Areolar  tissue,  action  of  acetic  acid  on,  22 

corpuscles,  22 

stained  with  logwood,  22 

stained  with  picrocarmine,  22 

constrictions  on  fibre-bundles,  23 

elastic  fibres,  21 

—  stained  with  magenta,  20 

stained  with  picrocarmine,  20 

interstitial  injection  of  osmic  acid,  22 

interstitial  injection  of  picric  acid,  22 

Arrector  pili  muscle,  93 
Arteries,  51 
Articular  cartilage,  14 
Artificial  digestion,  92 
Arthropoda,  muscles  of,  39 
Arytenoid  cartilage  of  ox,  18 
Auerbach's  plexus,  71 
Axis-cylinder  in  nerve-fibre,  42 


BRO 

BARYTA  water,  xxxiv 
Basilar  membrane,  1 14 
Berlin  blue,  li 
Bicarbonate  of  soda,  34 
Bichromate  of  ammonia,  xxxii 

—  potash,  xxxi 
Bile-ducts,  74 

Birch  on  bone  lamella,  32 
Bladder  of  frog,  36 

—  of  mammal,  89 
Blood,  circulation  of,  52 

—  granules  of,  6 
Blood-corpuscles  of  frog,  i 
of  newt,  I 

man,  5 

effect  of  acetic  acid,  2 

of  dilute  alcohol,  2 

of  ammonium  chromate,  3 

of  hydrochloric  acid,  i 

—  ■ of  magenta,  2 

of  osmic  acid,  3 

of  picric  acid,  3 

of  syrup,  2 

of  water,  2 

of  tannic  acid,  2 

feeding  of  colourless,  4 

structure  of,  3 

Blood-crystals,  6 
Blood-serum,  xxx 
Blood-vessels,  49 

—  development  of,  50 

—  injection  of,  Iv 
Bone,  cancellous,  34 

—  dense,  33 

—  foetal,  32 
Bone-corpuscles,  31 
Bone-lamelte  and  Sharpey's  fibres,  32 

—  methods  of  softening,  30 
Bowman's  glands,  1 1 1 

—  sarcous  elements,  39 
Brain,  loi 

Bronchus,  structure  of,  55 
Brownian  movement. 


126 


INDEX. 


CRU 


DUC 


Briicke's  dissecting  microscope,  xxxv 
Brunner's  glands,  68 


CABINET  for  microscopic  specimens,  xx 
Caliper  micrometer,  xxv 
Camera  lucida  of  Chevalier,  xxvii 

Zeiss,  xxvii 

Canada  balsam,  xlix 

Canaliculi,  31 

Cannulas  for  injecting,  Hi 

Capillaries,  49 

Carmine  staining-fluid,  xliii 

—  gelatine  injection,  li 
Carotid  gland,  82 

Carrot  for  imbedding,  xxxvi 
Cartilage,  articular,  fresh,  14 

—  cell-spaces  of,  14 

—  matrix,  14 
Cartilage-cells,  action  of  water,  14 

—  action  of  iodine,  14 

—  preservation  of,  14 

—  fat  in,  16 

—  stained  by  gold  chloride,  17 

—  stained  by  eosin,  16 

—  stained  by  silver  nitrate,  17 
Cartilage,  costal,  adult,  16 
young,  1 5 

—  transition  between  hyaline  and  yellow,  19 

—  hyaline,  14 

—  elastic,  18 

—  fibrous,  18 

—  of  Loligo,  ig 
Cell-spaces  of  tendon,  25 
Cells  of  tendon,  25 
Cements,  1 

Cement-substance  of  muscle,  37 
Central  tendon  of  diaphragm,  76 
Cerebellum,  100 

—  double-staining  of,  lOO 
Cerebrum,  loi 

Chalice  cells,  10 
Chloral  hydrate  for  brain,  96 
Chloride  of  gold,  xlv 
Chloroform  on  cilia,  12 
Choroid  of  eye,  107 
Chromate  of  ainmonia,  xxxii 
Chromic  acid,  xxxi 

and  spirit,  xxxi 

and  nitric  acid,  xxxiii 

Cilia,  1 1 

Ciliary  motion,  1 1 

on  gills  of  a  mussel,  1 1 

—  effect  of  heat,  1 1 

—  effect  of  chloroform,  1 2 
Ciliary  muscle,  108 
Ciliated  epithelium,  1 1 
Circulation  of  the  blood,  52 
Circumvallate  papillte,  60 


Cleaning  cover-glasses,  xxv 
Cleaning  slides,  xxiv 
Cleavage  of  muscular  fibres,  39 
Clove  oil,  XX 
Coccygeal  gland,  82 
Cochlea,  1 13 
Colostrum,  122 
Columnar  epithelium,  10 
Commissures  of  the  cord,  97 
Condenser,  bull's  eye,  xxi 
Cones  of  retina,  1 10 
Connective-tissue  corpuscles,  21 
Convoluted  tubules  of  kidney,  85 
Corium,  91 
Cornea,  cell-spaces  of,  106 

—  corpuscles  and  nerves  of,  105 

—  method  of  preparing,  105 
Corpora  lutea,  121 

Corti,  organ  of,  114 

—  rods  of,  114 

—  cells  of,  115 
Costal  cartilage,  1 5 
Cover-glasses,  xix 

—  measuring  of,  xxv 
Crusta  petrosa,  62 
Crystalline  lens,  107 
Cutis  vera,  structure  of,  90 
Cutting  sections,  methods  of,  xxxvi 


DAMMAR,  xlviii 
Decalcifying  solutions,  xxxiii 
DeciduK,  124 
Deiters,  cells  of,  115 
Delineation  of  objects,  xxvi 
Dentine,  62 
Diapedesis,  52 
Diaphragm,  of  microscope,  xxiii 

—  lymphatics  of,  76 

—  tendon  of,  41 

Digestion,  method  of  (W.  Stirling),  92 
Dilute  spirit,  xxxiv 
Dissecting  microscope,  xxxv 
Dissociation  by  needles,  xxxv 

—  by  teasing,  xxxv 

—  by  shaking,  29 
Dissociating  fluids,  xxxiv 
Distilled  water,  xlv 
Double-staining,  method  of,  xl 

—  cerebellum,  100 

—  bone,  35 

—  glands,  61 

—  Peyei^s  patch,  69 

—  retina,  1 10 

—  tendon,  25 

—  trachea,  54 
Duodenum,  67 
Ductless  glands,  82 
Ductus  cochleae,  114 


INDEX. 


127 


EAR 

pAR,  113 

J — '     Ebner,  v,,  method  for  bone,  xxxiii 
Elastic  cartilage,  iS 

lamina  of  cornea,  104 

Elastic  tissue,  20 

transverse  section  of,  21 

network  in  mesocolon,  2  r 

laminK  of  arteries,  50 

Elder-pith,  xxxvi 
Embryonal  tissue,  28 
Enamel,  62 
Endocardium,  47 
— ■  fibres  of  Purkinje,  48 
End-bulbs  of  penis,  1 19 
End-plates,  45 
Endomysium,  37 
Endothelium,  13 
Eosin,  xiv 
Epidermis,  90 
Epididymis,  117 
Epiglottis,  18 
Epineurium,  44 
Epithelium,  single  layer,  13 

—  squamous  stratified,  8 

—  ciliated,  1 1 

—  columnar,  10 

—  transitional,  12 

—  secretor)',  12 

Examining  a  simple  object,  xxiv 
Eye,  104 
Eye-lids,  1 10 
Eyepiece,  xxii 

FALLOPIAN    TUBE,  121 
Farrant's  solution,  xlviii 
Fat-cells,  27 

—  membrane  of,  27 

—  development  of,  27 
Feeding  blood-corpuscles,  4 
Ferrein,  pyramid  of,  85 
Fibrilte  of  muscle,  39 

Fibrils,  intra-cellular  plexus  of,  9 

—  intra-nuclear  plexus  of,  9 
Fibrin  in  blood,  5 
Fibro-cartilage,  14 
Fibrous  tissue,  white,  21 

yellow,  20 

Filiform  papilUi;,  60 
Forceps,  xix 

Formic  acid,  xlv 

Freezing  microtomes,  xxxviii 

—  mixture,  xxxviii 

—  process,  xxxviii 

Fresh  tissues,  examination  of,  xxx 
Fungiform  papillse,  60 


G 


ANGLIA,  of  bladder,  89 
—  lung,  59 


HYD 

Ganglia,  of  tongue,  61 

—  Gasserian,  102 

—  spinal,  102 

—  sympathetic,  103 
Gas-chamber,  xxix 
Gases,  application  of,  xxix 
Gasserian  ganglion,  103 
Gastric  follicles,  67 

—  method  of  isolating,  67 
Gauge  for  cover-glasses,  xxv 
Gelatine  injections,  li 
Germinal  vesicle,  120 
Giant-cells,  35 

Glass  slides,  xix 
Glandular  epithelium,  12 
Glisson's  capsule,  T>, 
Glycerine,  xlviii 
Glycerine  jelly,  xlviii 
Goblet-cells,  10 
Gold  chloride,  xlv 

—  for  Auerbach's  plexus,  71 
• —  cornea,  105 

—  nerve-terminations,  45 

—  skin,  95 

—  and  formic  acid,  xlv 
Graaffian  follicle,  120 
Granular  layer  of  cerebellum,  100 
• muscle,  40 

retina,  109 

Granules,  free  in  blood,  5 
Grey  matter  of  cerebrum,  !0[ 

of  cord,  97 

Gum  for  imbedding,  xxxviii 

HEMACYTOMETER,  7 
Hasmaglobinometer,  7 
Hjematoxylin,  xlii 
Haimin,  7 

Haemoglobin  crystals,  6 
Hairs,  94 
Hair-follicle,  93 
Hand  sections,  94 
Hardening,  methods  of,  xxx 
Hartnack's  microscope,  xxi 
Hassall's  corpuscles,  8t 
Haversian  canals,  31 
Heart,  muscular  substance  of,  47 

—  frog  and  mammal,  47 

Heat,  methods  of  applying,  xxviii 
Henle,  looped  tubules  of,  84 
Hensen,  cells  of,  115 
Hepatic  cells,  73 

—  vein,  -j^, 
High  powers,  xxii 
Hollow  ground  razor,  xix 
Hot  stage,  xxviii 
Hyaline  cartilage,  14 
Hydrochloric  acid  for  bone,  xxxiii 

—  —  blood,  1 


128 


INDEX. 


HYD 

Hydrochloric  acid  for  uriniferous  tubules,  87 
Hypophysis  cerebri,  82 

T  LLUMINATION,  methods  of,  xxiii 
-L     Imbedding,  methods  of,  xxxvi 
Incisures  of  a  nerve-fibre,  42 
Infundibulum  of  lung,  57 
Injection,  method  of,  i 

—  apparatus,  water  pressure,  Iv 

—  apparatus,  mercury  pressure,  liv 

—  of  an  entire  animal,  Iv 

—  of  blood  vessels,  Iv 

—  of  lymphatics,  Iv 

—  mass,  method  of  preparing,  li 
Intervertebral  disc,  18 
Intramembranous  bone,  35 
Intercellular  substance,  13 
Intestine,  68 

Intra-cellular  network,  9 
Intra-nuclear  network,  9 
Involuntary  muscle,  36 
Iodine-green,  xlvii 
Iodine  solution  for  cartilage,  14 
Iodised  serum,  xxxiv 
Irrigation,  2 

KIDNEY,  structure  of,  84 
Kleinenberg's  logwood,  xliii 

—  picric  acid,  xxxii 
Krause's  membrane,  39 

LABELS,  xxvi 
J     Lachrymal  gland,  no 
Lamp,  xxiii 
Large  intestine,  70 

—  slides,  xix 
Larynx,  53 
Lateral  discs,  39 
Lens,  107 

Lieberkiihn's  follicle,  70 
Ligamentum  nuchas,  20 
Lip,  60 

List  of  apparatus,  xix 

staining  reagents,  xlvii 

histological  requisites,  xix 

Liver,  71 

Living  tissues,  study  of,  xxvii 

Logwood  staining-fluids,  xlii 

Lung,  53 

Lymphatics,  injection  of,  Iv 

—  of  diaphragm,  76 
Lymph-glands,  simple,  78 
compound,  77 

MAGENTA,  xliv 
—  for  elastic  fibres,  20 

—  for  blood-corpuscles,  xliv 
Malassez,  7 


NER 
Malpighian  bodies,  79 

—  capsules,  84 

—  tufts,  84 

Mammary  glands,  122 
Marrow,  red,  35 

—  giant-cells  of,  35 
Measuring  cover-glasses,  xxv 
Medulla  oblongata,  99 
Medullated  nerve-fibres,  42 
Membrana  granulosa,  120 
Meissner's  plexus,  71 
Mesentery  of  newt,  38 
Mesocolon  of  rabbit,  2 1 
Methyl-aniline,  xliv 
Microscope,  xxi 

—  compound,  xxi 

—  dissecting,  xxxv 

—  preparation  of,  xxi 

—  testing  of,  xxi 

—  Hartnack's,  xxi 

—  Swift's,  xxi 
Microtomes,  xxxvii 

■ —  Ranvier's  hand,  xxxvii 

—  A.  B.  Stirling's,  xxxvii 

—  Rutherford's  freezing,  xxxviii 

—  Williams'  freezing,  xxxix 

—  B.  Lewis's  ether,  xl 
Milk,  122 
Modiolus,  114 
Moist  chambers,  xxvii 
Mounting  of  specimens,  1 

—  solutions,  xlviii 
Mouth,  60 
Mucigen,  64 
Mucin,  8 

Muco-salivary  glands,  64 
Mucous  glands  of  tongue,  61 
Mucous  tissue,  28 
Miiller's  fibres,  109 

—  fluid,  xxxii 
Multipolar  nerve-cells,  98 
Muscle,  non-striped,  36 

—  striped,  38 

—  and  tendon,  union  of  41 

—  of  crab,  39 

—  of  heart,  47 

—  nerves  of,  45 
Muscularis  mucosEe,  66 

NAILS,  94 
Nasal  mucous  membrane   1 1 
Needles  mounted,  xix 
Nerve-cells  of  ganglia,  102 

spinal  cord,  98 

frog,  103 

Nerve-fibres,  medullated,  42 

—  non-medullated,  45 

—  of  spinal  cord,  44 

—  with  osmic  acid,  43 


INDEX. 


129 


NER 

Nerve-fibres,  silver  nitrate,  43 

—  of  cornea,  105 

muscle,  45 

skin,  95 

—  terminal  organs,  45 
Nerve-fibrils,  105 
Neurilemma,  44 
Neurogleia,  97 

Neutral  tint-reflector,  xxvii 
Nodes  of  Ranvier,  43 
Non-striped  muscle,  36 
Nosepiece,  xxii 

OBJECTIVES,  xxi 
—  dry  and  immersion,  xxii 
Ocular,  xxii 
Odontoblasts,  63 
Gisophagus,  65 
Oil  of  cloves,  XX 
Oil  immersion  lenses,  xxii 
Olfactory  cells,  1 1 1 
Olivary  body,  99 
Omentum,  26 
Opaque  injections,  74 
Osmic  acid,  xlvi 
Ossification,  intra-cartilaginous,  34 

—  intra-membranous,  35 
Osteoblasts,  33 
Osteoclasts,  35 

Outline  plates,  how  to  fill  in,  xvii 

Ovary,  120 

Ova,  development  of,  121 

PACINIAN  corpuscles,  46 
Pancreas,  75 
Papillje  of  tongue,  60 
Paraffine  for  imbedding,  xxxvi 
Parotid  gland,  64 
Penis,  119 
Peptic  glands,  67 
Peptic  cells,  66 
Pericardium,  47 
Perimysium,  37 
Perineurium,  44 
Periosteum,  31 

Peripheral  terminations  of  nerves,  45 
Peyer's  glands,  69 
Photophore,  xxxv 
Pia  mater,  vessels  of,  49 
Picric  acid,  xxxii 

Kleinenberg's,  xxxii 

for  bone,  30 

Picrocarminate  of  ammonia,  xliii 
Pigment-cells  of  choroid,  107 
Pith  for  imbedding,  xxxvi 
Placenta,  123 
Plasma  cells,  22 
Pleura,  56 


SER 


Potassic  bichromate,  xxxi 

Potassic  hydrate,  xxxiv 

Pressure  bottle,  injection  apparatus,  liv 

Prickle  cells,  60,  91 

Prism  for  drawing,  xxvii 

Prostate,  119 

Prussian  blue,  soluble,  li 

Pulmonary  vessels,  injection  of,  58 

Pulp  of  tooth,  62 

spleen,  79 

Pure  soluble  blue,  li 
Purkinje's  fibres  of  heart,  48 
■ —  cells  of  cerebellum,  100 
Purpurine,  xliv 
Pyloric  end  of  stomach,  67 

RANVIER'S  microtome,  xxxvii 
—  nodes,  43 

—  photophore,  xxxv 

—  picrocarmine,  xliii 
Razor,  xix 
Reagents,  bottles  for,  xx 

—  list  required,  xx 
Reissner,  membrane  of,  114 
Respiratory  organs,  53 
Rete  Malpighii,  90 
Retina,  108 

Rodded  epithelium  of  kidney,  86,  87 

Rods  of  retina,  109 

Rolando  substantia  gelatinosa,  97 

Rosanilin  nitrate,  xliv 

Rosein,  xlv 

SALINE  solution,  xxx 
Saliva,  8 
Salivary  corpuscles,  8 

—  glands,  64 
Salt  solution,  xxx 
Sarcolemma,  38 
Sarcous  elements,  39 
Scala  vestibuli,  114 
Scalp,  93 

Schafer's  heating  apparatus,  xxviii 
Schneiderian  mucous  membrane,  1 1 1 
Scissors,  xix 
Schwann,  sheath  of,  42 

—  white  substance  of,  43 
Sclerotic,  107 

Sealing  preparations,  1 
Sebaceous  glands,  93 
Sections,  making  of,  xxxvi 

—  mounting  of,  xlix 

—  how  to  float  on  to  a  slide,  xxvi 
Semicircular  canals,  113 
Seminiferous  tubules,  116 

Septum  cysterniE  lymphaticoe  maguK,  76 
Serous  membranes,  13 

—  glands,  61 


I30 

SHA 

Sharpey's  fibres,  32 
Silver  nitrate,  xlv 

—  lines,  13 

—  process,  xlv 
Skin,  90 

—  blood-vessels  of,  94 

—  nerves  of,  95 
Slides,  xix 
Small  intestine,  67 
Softening  fluids,  xxxiii 
Solitary  glands,  70 
Spermatoblasts,  117 
Spermatozoa,  1 1 8 
Spinal  cord,  97 

isolation  of  cells  of,  98 

Spinal  ganglia,  102 
Spiral  groove,  114 

—  lamina,  1 14 

—  ligament,  114 
Spleen,  78 

Squamous  epithelium,  8 
Stage  of  microscope,  xxii 

—  warm,  xxviii 
Staining  fluids,  xlvii 

—  methods  of,  xlii 

Stirling,  W.,  method  of  artificial  digestion,  92 
Stomach,  66 
Stomata,  76 
Stratified  epithelium,  g 
Stratum  corneum,  90 

—  lucidum,  90 

—  granulosum,  90 

—  Malpighii,  90 
Strieker's  warm  stage,  xxviii 
Striped  muscle,  38 
Sub-arachnoid  trabecule,  23 
Sub-lingual  gland,  64 
Sub-maxillary  gland,  64 
Supra-renal  bodies,  82 
Sweat-glands,  91 
Sympathetic  nerves,  45 
Syringe  for  injecting,  Hi 

—  subcutaneous,  22 

TACTILE  corpuscles,  92 
Tadpole,  tail  of,  50 
Tail  of  a  rat,  25 

Tannic  acid,  effect  of,  on  blood,  2 
Taste-bulbs,  62 
Teasing,  method  of,  xxxv 
Tectorial  membrane,  1 14 
Tendon,  23 

—  cells,  25 
Test  objects,  xxi 
Testis,  116 

—  of  newt,  118 
Thin  slides,  xix 


INDEX. 


ZON 


Thymus  gland,  81 
Thyroid  gland,  82 
Tongue,  60 

—  mucous  glands  of,  61 

—  serous  glands  of,  61 
Tonsils,  65 

Tooth,  62 
Trachea,  54 

Transitional  epithelium,  12 
Treble-staining,  xlvii 
Trypsin  digestion,  xxxiv 
Tubuli  seminiferi,  84 
Tunica  adventitia,  49 

—  adnata,  1 16 

—  albuginea,  1 16 

—  granulosa,  1 20 
Turntable,  xlix 

UMBILICAL  cord,  124 
,     Unstriped  muscle,  36 
Ureter,  88 
Urinary  bladder,  89 
Uriniferous  tubules,  84 
Uterus,  121 

VAS  deferens,  117 
Vasa  efterentia  of  testis,  31 
Vasa  recta,  kidney,  88 

testis,  116 

Veins,  51 

Villi  of  small  intestine,  68 
—  of  placenta,  123 
Vitelline  membrane,  120 
Vitreous  humour,  28 
Vocal  cords,  53 
Voluntary  muscle,  38 

WAGNER'S  corpuscles,  92 
Warm  stages,  xxviii 
Water,  action  of,  on  blood-corpuscles,  2 
Wharton's  jelly,  28 
White  blood-corpuscles,  3 

method  of  feeding,  4 

White  fibro-cartilage,  18 

White  fibrous  tissue,  21 

White  matter  of  cerebellum,  100 

of  cerebrum,  loi 

of  spinal  cord,  97 

White  substance  of  Schwann,  42 

'WELLOW  fibro-cartilage,  18 

ZEISS,  camera  lucida,  xxvi 
—  oil  immersion  lenses,  xxi 
Zinc  cement,  1 
Zona  pellucida,  120 


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A    MANUAL    of    MIDWIFERY     for     MIDWIVES.      By 

Fancoubt  Barnes,  M.D.  Aber.,  M.R.C.P.  Lond.,  Physician  to  the 
British  Lying-in  Hospital;  Assistant  Physician  to  the  Royal  Maternity 
Chanty  of  London  ;  Physician  for  the  Diseases  of  Women  to  the  St. 
George's  and  St.  James's  Dispensary.  Crown  Svo.  with  numerous 
Illustrations,  6s. 

A  TREATISE  on  the  THEORY  and  PRACTICE  of  MEDI- 
CINE. By  John  Syer  Bristowe,  M.D.  Lond.,  Fellow  and  formerly 
Censor  of  the  Royal  College  of  Physicians  ;  Senior  Physician  to,  and 
Joint  Lecturer  on  Medicine  at,  St.  Thomas's  Ho'^pital  ;  President  of 
the  Society  of  Medical  Officers  of  Health  ;  Examin<.-r  in  Medicine  to 
the  Royal  College  of  Surgeons,  formerly  Examiner  in  Medicine  to  the 
University  of  London,  and  Lecturer  on  Genera!  Pathology  and  on 
Physiology  at  St.  Thomas's  Hospital.  Third  Edition,  revised  and 
enlarged,  price  21s. 

The  FUNCTIONS  of  the  BRAIN.  With  numerous  Illustra- 
tions. By  David  Ferrier,  M.D.,  F  R.S.,  Assistant  Physirian  to 
King's  College  Hospital  ;  Professor  of  Forensic  Medicine,  King's 
College.      8vo.  15^. 

The   LOCALISATION   of  CEREBRAL   DISEASES.     By 

David  Ferrier,  M.D.,  F.R.S.  With  numerous  Illustrations.  Svo. 
JS.  6d. 

A  GUIDE  to  THERAPEUTICS.     By  Robert  Farquhar- 

soN,  M.P.,  M.D  Edin.,  F.R.C.P.  Lond.,  late  Lecturer  on  Materia 
Medica  at  Si.  Mary's  Hospita  1  Medical  School,  &c.  Second  Edition. 
Crown  Svo.  js.  6d. 

An  EPITOME  of  THERAPEUTICS.  Being  a  Comprehen- 
sive Summary  of  the  I'reatment  of  Disease  as  recommended  by  the 
leading  British,  American,  and  Continental  Physicians.  By  W. 
Domett  Stone,  M.D.,  F.R.C.S.,  Honorary  Member  of  the  College 
of  Physicians  of  Sweden,  Physician  to  the  Westminster  General 
Dispensary;  Editor  of  the  'Half-yearly  Abstract  of  the  Medical 
Sciences.'     Crown  Svo.  Sj.  6d. 


London:  SMITH,  ELDER,  &  CO.,   15  Waterloo  Place. 


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